Methods for making and using endoxifen

ABSTRACT

The present disclosure provides industrially scalable methods of making (Z)-endoxifen or a salt thereof, crystalline forms of endoxifen, and compositions comprising them. The present disclosure also provides methods for treating hormone-dependent breast and hormone-dependent reproductive tract disorders.

CROSS-REFERENCE

This application is a U.S. National Phase Application under 35 U.S.C. §371 of International Application No. PCT/US2018/050272, filed Sep. 10,2018, which claims the benefit of U.S. Provisional Application No.62/556,799, filed Sep. 11, 2017; U.S. Provisional Application No.62/556,884, filed Sep. 11, 2017; U.S. Provisional Application No.62/624,787, filed Jan. 31, 2018; and U.S. Provisional Application No.62/693,885, filed Jul. 3, 2018, each incorporated herein by reference intheir entireties.

BACKGROUND OF THE INVENTION

Breast cancer is by far the most common form of cancer in women, and itis the second leading cause of cancer death in humans. Despite advancesin diagnosing and treating breast cancer, the prevalence of this diseasehas been steadily rising at a rate of about 1% per year since 1940.Today, the likelihood that a woman living in North America will developbreast cancer during her lifetime is one in eight. In addition to breastcancers, other breast disorders affecting a large number of womeninclude benign but often precancerous lesions, such as ductalhyperplasia, lobular hyperplasia, atypical ductal hyperplasia, andatypical lobular hyperplasia.

Current best practice for the treatment of breast cancer is to diagnosebreast cancer with mammography and then treat the patient with surgery,radiation therapy, and chemotherapy. The current widespread use ofmammography has resulted in improved detection of breast cancer.Although breast cancer is about 100 times less common among men thanamong women, the lifetime risk of men getting breast cancer is about 1in 1,000. About 2,470 new cases of invasive breast cancer will bediagnosed in men and about 460 men will die from breast cancer. AmericanCancer Society estimates that in 2017, 255,180 new cases of breastcancer will be diagnosed in both men and women combined and 41,070subjects will die of breast cancer. Nonetheless, the death rate due tobreast cancer has remained relatively unchanged at about 21 deaths per100,000 women and 0.4 deaths per 100,000 men. All too often, breastcancer is discovered at a stage that is too far advanced, whentherapeutic options and survival rates are severely limited.

Breast cancers include any malignant tumor of breast cells. There areseveral types of breast cancer. Exemplary breast cancers include, butare not limited to, ductal carcinoma in situ, lobular carcinoma in situ,invasive (or infiltrating) ductal carcinoma, invasive (or infiltrating)lobular carcinoma, inflammatory breast cancer, triple-negative breastcancer, ER+ breast cancer, HER2+ breast cancer, adenoid cystic (oradenocystic) carcinoma, low-grade adenosquamous carcinoma, medullarycarcinoma, mucinous (or colloid) carcinoma, papillary carcinoma, tubularcarcinoma, metaplastic carcinoma, and micropapillary carcinoma. A singlebreast tumor can be a combination of these types or be a mixture ofinvasive and in situ cancer.

Tamoxifen is a selective estrogen receptor modulator that is used forthe treatment of women with endocrine responsive breast cancer, i.e.,hormone-dependent or hormone-sensitive breast cancer. Adjuvant therapyprimarily via oral delivery of tamoxifen is known to have severe sideeffects such as vasomotor symptoms, for example hot flashes, andreproductive tract (gynecologic) cancers. Patient compliance remains aproblem with tamoxifen therapy. Further, the majority of the individualson adjuvant tamoxifen therapy do not respond to the drug and 30-50% ofthe patients subsequently die of their disease.

Several cytochrome P450 (CYP) mutations have been proposed to causereduced conversion of tamoxifen to its active metabolite, endoxifen, andreduce tamoxifen efficacy and increase resistance to the drug (Dickschenet al. Front Pharmacol. 2012; 3: 92. PMCID: PMC3357105). So far, over140 allelic variants of CYP2D6 have been described and a substantialpart of these are associated with reduced or absent activity of theencoded enzyme. Based on the combination of the carried alleles, eachindividual subject can be classified into one of four phenotypic groups:poor metabolizer (PM) with plasma endoxifen levels less than 30 nM,intermediate metabolizer (IM), extensive metabolizer (EM) andultra-rapid metabolizer (UM), reflecting variations in levels of serumendoxifen. However, changes in the CYP genotype do not fully explain thetamoxifen resistance and the reduced endoxifen levels observed in somesubjects.

Therefore, several alternatives to tamoxifen are being developed for thetreatment of breast cancer, which include low dose-tamoxifen (LazzeroniM et al. Breast Cancer Res. 2012 Oct. 29; 14(5):214) and tamoxifen'sactive metabolites, afimoxifene (see, U.S. Pat. Nos. 7,485,623;7,507,769; 7,704,516; 7,786,172; 7,968,532; and 8,048,927; Mansel R etal. Breast Cancer Res Treat. 2007 December; 106(3):389-97. PubMed PMID:17351746; Rouanet P et al. J ClinOncol. 2005 May 1; 23(13):2980-7.PubMed PMID: 15860853), endoxifen (see, U.S. Pat. Nos. 9,333,190;9,220,680; 9,090,640; and 9,200,045; U.S. Publication Nos. 2009/0291134and US20100112041), and their derivatives (see, U.S. Pat. No. 8,063,249;U.S. Publication Nos. 2015/0080339 and 2014/0193334). It is widelyaccepted that (Z)-endoxifen is the main active metabolite responsiblefor the clinical efficacy of tamoxifen.

While hydrochloride and citrate salts of endoxifen (See, e.g., Fauq etat., Bioorganic & Medicinal Chemistry Letters. 20 (2010) 3036-3038;Stearns et al., J. Natl. Cancer Inst. Vol 95, No. 23, 2003; USPublication Nos. 2009/0291134 and 2010/0112041; Clinical Trials Gov.Identifier Nos. NCT01273168 and NCT02311933; Goetz et al., 2015, SanAntonio Cancer Symposium; Ahmad et al., Clinical Pharmacology &Therapeutics. 88(6) 814-817, 2010; and J Clin. Oncol. 30, 2012 (suppl;abstr 3089); Ahmad et al. Breast Cancer Research and Treatment 2010,122, 579-584) are known in the art and currently under evaluation formetastatic cancer, there remains unmet medical need for new compositionsand methods for the treatment and/or prevention of hormone-dependentbreast and reproductive tract (gynecologic) disorders.

SUMMARY OF THE INVENTION

The present disclosure addresses this need by providing compositions andmethods for the treatment and/or prevention of hormone-dependent breastand reproductive tract (gynecologic) disorders. In certain aspects, thepresent disclosure provides novel industrially scalable methods ofmaking Z-endoxifen or salts thereof, crystalline forms of endoxifen, andcompositions comprising them. In certain aspects, the present disclosureprovides novel crystalline forms of endoxifen which may provideadvantages including improved bioavailability and stability relative toother crystalline or amorphous forms.

In certain aspects, the present disclosure provides a compositioncomprising a crystalline form of a compound of Formula (III):

In some embodiments, at least 90% by weight of the compound of Formula(III) in the composition is the (Z)-isomer. In some embodiments, thecrystalline form is Form I of the compound of Formula (III). CrystallineForm I may be characterized by an x-ray powder diffraction patterncomprising major peaks at 16.8±0.3°, 17.1±0.3° and 21.8±0.3° two theta,and optionally further comprising at least one peak selected from16.0±0.3°, 18.8±0.3° and 26.5±0.3° two theta. In some embodiments, thex-ray powder diffraction pattern further comprises at least one peakselected from 12.3 0.3°, 28.0±0.3° and 29.0±0.3° two theta. The x-raypowder diffraction pattern may further comprise peaks at 12.3±0.3°,16.0±0.3°, 18.8±0.3°, 26.5±0.3°, 28.0±0.3° and 29.0±0.3° two theta. Insome embodiments, crystalline Form I is characterized by an x-ray powderdiffraction pattern substantially as set forth in FIG. 9 or FIG. 10.Greater than 90%, 95% or 99% by weight of the compound of Formula (III)in the composition may be crystalline Form I. In some embodiments, thecomposition comprises 0.01 mg to 200 mg of crystalline Form I, such asabout 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of crystalline Form I.

In some embodiments, the composition comprises the (E)-isomer and the(Z)-isomer of the compound of Formula (III) in an E/Z ratio between 0.9and 1.3, such as about 1.1. In some embodiments, the crystalline form isForm II of the compound of Formula (III). Crystalline Form II may becharacterized by an x-ray powder diffraction pattern comprising majorpeaks at 7.0±0.3°, 11.9±0.3°, 14.0±0.3° and 18.4±0.3° two theta, andoptionally further comprising a peak at 22.0±0.3° two theta. In someembodiments, the x-ray powder diffraction pattern further comprises atleast one peak selected from 6.6±0.3°, 13.3±0.3° and 20.0±0.3° twotheta. The x-ray powder diffraction pattern may further comprise peaksat 6.6±0.3°, 13.3±0.3°, 20.0±0.3° and 22.0±0.3° two theta. In someembodiments, crystalline Form II is characterized by an x-ray powderdiffraction pattern substantially as set forth in FIG. 11 or FIG. 12.Greater than 90%, 95% or 99% by weight of the compound of Formula (III)in the composition may be crystalline Form II. In some embodiments, thecomposition comprises 0.01 mg to 200 mg of crystalline Form II, such asabout 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of crystalline Form II.

In some embodiments, the composition comprises the (E)-isomer and the(Z)-isomer of the compound of Formula (III) in an E/Z ratio between 0.9and 1.3, such as about 1.1. In some embodiments, the crystalline form isForm III of the compound of Formula (III). Crystalline Form III may becharacterized by an x-ray powder diffraction pattern comprising majorpeaks at 11.9±0.3°, 13.9±0.3°, 17.1±0.3° and 17.7±0.3° two theta, andoptionally further comprising a peak at 25.3±0.3° two theta. In someembodiments, the x-ray powder diffraction pattern further comprises atleast one peak selected from 18.2±0.3°, 22.5 0.3° and 26.8±0.3° twotheta. The x-ray powder diffraction pattern may further comprise peaksat 18.2±0.3°, 22.5±0.3°, 25.3±0.3° and 26.8±0.3° two theta. In someembodiments, crystalline Form III is characterized by an x-ray powderdiffraction pattern substantially as set forth in FIG. 13. Greater than90%, 95% or 99% by weight of the compound of Formula (III) in thecomposition may be crystalline Form III. In some embodiments, thecomposition comprises 0.01 mg to 200 mg of crystalline Form III, such asabout 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of crystalline Form III.

In certain aspects, the present disclosure provides a pharmaceuticalcomposition comprising a pharmaceutically acceptable carrier or diluentand the composition comprising a crystalline form of a compound ofFormula (III). A composition of the present disclosure may be formulatedfor oral, parenteral, topical, or intraductal delivery. In someembodiments, the composition is formulated for oral delivery as atablet, a caplet, a capsule, or a pill. In some embodiments, a meanhalf-life of endoxifen in a subject treated with the composition isbetween 30 hours to 60 hours. A composition comprising a crystallineform of a compound of Formula (III) may be formulated for oral deliveryas an enteric tablet, an enteric caplet, an enteric capsule, adelayed-release tablet, a delayed-release caplet or a delayed-releasecapsule. In some embodiments, the composition is administered to asubject for the treatment or prevention of a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both inthe subject.

In certain aspects, the present disclosure provides an oral compositioncomprising 1 mg to 200 mg per unit dose of the composition comprising acrystalline form of a compound of Formula (III) for administration to asubject in need thereof, wherein daily administration of the oralcomposition achieves in the subject:

a steady state plasma level of endoxifen within 7 to 21 days;

a steady state plasma level of endoxifen ranging from 25 nM to 300 nM;

a steady state plasma level of endoxifen greater than 30 nM;

maximal plasma levels of endoxifen within 2 to 10 hours afteradministering; or

any combination thereof.

In some embodiments, a mean half-life of endoxifen in a subject treatedwith the composition comprising a crystalline form of a compound ofFormula (III) is between 40 hours to 55 hours. In some embodiments, thecomposition is formulated as an enteric tablet, an enteric caplet, anenteric capsule, a delayed-release tablet, a delayed-release caplet or adelayed-release capsule. In some embodiments, at least 40%, at least50%, at least 60%, at least 70%, at least 80%, or at least 90% ofendoxifen in the composition is released in the intestines. In someembodiments, the composition comprising a crystalline form of a compoundof Formula (III) exhibits a mean area under the curve extrapolated totime infinity (AUC_(0-inf)) of endoxifen of 200 hr*ng/mL to 10000hr*ng/mL, of 300 hr*ng/mL to 8000 hr*ng/mL, of 400 hr*ng/mL to 6000hr*ng/mL or of 700 hr*ng/mL to 6000 hr*ng/mL.

In certain aspects, the present disclosure provides a method of treatinga subject in need thereof, the method comprising administering to thesubject the composition comprising a crystalline form of a compound ofFormula (III). In some embodiments, the subject has or is at risk ofhaving a hormone-dependent breast disorder, a hormone-dependentreproductive tract disorder, or both. The hormone-dependent breastdisorder or the hormone-dependent reproductive tract disorder may be abenign breast disorder, hyperplasia, atypia, atypical ductalhyperplasia, atypical lobular hyperplasia, increased breast density,gynecomastia, DCIS, LCIS, breast cancer, precocious puberty,McCune-Albright Syndrome, endometrial cancer, ovarian cancer, uterinecancer, cervical cancer, vaginal cancer, or vulvar cancer. In someembodiments, the subject has prostate cancer and has or is at risk ofhaving gynecomastia. The subject may have tamoxifen-refractory ortamoxifen resistant hormone-dependent breast disorder orhormone-dependent reproductive tract disorder. In some embodiments, thesubject is or will be treated with an SSRI drug selected from the groupconsisting of citalopram, escitalopram, fluoxetine, paroxetine,sertraline, and vilazodone.

In practicing any of the subject methods, the composition may comprise0.01 mg to 200 mg of (Z)-endoxifen. In some embodiments, the subject isadministered about 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of(Z)-endoxifen daily. In some embodiments, a steady state plasma level ofendoxifen in the subject is greater than 30 nM. The steady state plasmalevel of endoxifen may be achieved within 7 to 21 days of the firstadministration of the composition. In some embodiments, time to maximumplasma levels of endoxifen ranges from 2 hours to 10 hours or from 4hours to 8 hours after administering the composition.

In certain aspects, the present disclosure provides a method of treatinga subject having or at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both, themethod comprising administering the composition comprising a crystallineform of a compound of Formula (III), wherein administration of thecomposition achieves:

a mean half-life of endoxifen in the subject ranging from 30 hours to 60hours after administration;

a time to maximum plasma levels of endoxifen ranging from 4 hours to 8hours after administration; and

a steady state plasma level of endoxifen greater than 30 nM.

In some embodiments, the hormone-dependent breast disorder and thehormone-dependent reproductive tract disorder are selected from thegroup consisting of benign breast disorders, hyperplasia, atypia,atypical ductal hyperplasia, atypical lobular hyperplasia, increasedbreast density, gynecomastia, DCIS, LCIS, breast cancer, precociouspuberty, McCune-Albright Syndrome, endometrial cancer, ovarian cancer,uterine cancer, cervical cancer, vaginal cancer, and vulvar cancer.

In practicing any of the subject methods, the composition may comprise0.01 mg to 200 mg of (Z)-endoxifen. In some embodiments, the subject isadministered about 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of(Z)-endoxifen. The mean area under the curve extrapolated to timeinfinity (AUC_(0-inf)) of endoxifen may be 200 hr*ng/mL to 10000hr*ng/mL, 300 hr*ng/mL to 8000 hr*ng/mL, 400 hr*ng/mL to 6000 hr*ng/mLor 700 hr*ng/mL to 6000 hr*ng/mL. In some embodiments, the compositionis formulated as an enteric tablet, an enteric caplet, an entericcapsule, a delayed-release tablet, a delayed-release caplet or adelayed-release capsule. In some embodiments, at least 40%, at least50%, at least 60%, at least 70%, at least 80%, or at least 90% ofendoxifen in the composition is released in the intestines. Thecomposition may be administered once a day, twice a day, thrice a day,four times a day, every other day, twice a week, weekly, fortnightly,twice a month, monthly, quarterly, once every six months, or annually.

In one aspect, the present disclosure provides an industrially scalableprocess for manufacturing (Z)-endoxifen or salts thereof, comprising thesteps of: (a) subjecting a mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III), represented by

to fractional crystallization from a first solvent to form a firstcrystalline solid and first crystalline mother liquor, wherein the firstmother liquor has an E/Z ratio of at least 50% higher (Z) endoxifen ascompared with the E/Z ratio of the mixture of (E)-endoxifen and(Z)-endoxifen; (b) subjecting the first mother liquor torecrystallization from a second solvent by concentrating the firstmother liquor or by swapping out the first solvent from the first motherliquor one or more times with the second solvent, to form a secondcrystalline solid and a second mother liquor, wherein the secondcrystalline solid is ≥90% (Z)-endoxifen; and (c) optionally, subjectingthe second crystalline solid to recrystallization from a third solventor chromatographic treatment one or more times to form a thirdcrystalline solid. In some embodiments, any one or more of the firstsolvent, the second solvent, and the third solvent are preheated to atemperature ranging from 40° C. to 80° C.

In some aspects, the present disclosure provides an industriallyscalable process for manufacturing crystalline Form I of the compound ofFormula (III), comprising the steps of (a) subjecting a mixture of(E)-endoxifen and (Z)-endoxifen, compounds of Formula (III), representedby

to fractional crystallization from a first solvent to form a firstcrystalline solid and first mother liquor, wherein the first motherliquor has an E/Z ratio of at least 50% higher (Z) endoxifen as comparedwith the E/Z ratio of the mixture of (E)-endoxifen and (Z)-endoxifen;(b) subjecting the first mother liquor to recrystallization from asecond solvent by concentrating the first mother liquor or by swappingout the first solvent from the first mother liquor with the secondsolvent one or more times, to form a second crystalline solid and asecond mother liquor, wherein the second crystalline solid is ≥90%(Z)-endoxifen; and (c) subjecting the second crystalline solid torecrystallization from a third solvent or chromatographic treatment oneor more times to form a third crystalline solid, wherein the thirdcrystalline solid is crystalline Form I of the compound of Formula(III).

In certain aspects, the present disclosure provides a crystalline formof a compound of Formula (III) produced according to a method describedherein. In some embodiments, the crystalline form is Form I of thecompound of Formula (III). In some embodiments, the crystalline form isForm II of the compound of Formula (III). In some embodiments, thecrystalline form is Form III of the compound of Formula (III).

In another aspect, the present disclosure provides that the mixture of(E)-endoxifen and (Z)-endoxifen, compounds of Formula (III), in step ais pretreated with 6N HCl and neutralized with 8N NaOH.

In another aspect, the industrially scalable process further comprisespreparing the mixture of (E)-endoxifen and (Z)-endoxifen by coupling thecompound of Formula (II),(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl) methanone, topropiophenone mediated by a McMurry reaction via a titanium salt and areducing agent in an inert organic solvent to form the mixture of(E)-endoxifen and (Z)-endoxifen; and wherein the compound of Formula(II) has a structure represented by

In yet another aspect, the industrially scalable process furthercomprises preparing the compound of Formula (II) by demethylating[4-[2-(dimethylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone, thecompound of Formula (I), wherein the compound of Formula (I) has thestructure

with a demethylating agent and a proton acceptor in an inert organicsolvent to form the compound of Formula (II).

The present disclosure also provides an industrially scalable processfor manufacturing (Z)-endoxifen and salts thereof, comprising the stepsof: (a) subjecting a mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III) represented by

to fractional crystallization from ethyl acetate to form a firstcrystalline solid and first crystalline mother liquor, wherein the firstmother liquor has an E/Z ratio of at least 30% higher (Z) endoxifen ascompared with the E/Z ratio of the mixture of (E)-endoxifen and(Z)-endoxifen; (b) subjecting the first mother liquor torecrystallization from IPA or IPA/PPW (1:1 v/v) by concentrating thefirst mother liquor, or by swapping out ethyl acetate from the firstmother liquor one or more times with IPA or IPA/PPW, to form a secondcrystalline solid and a second mother liquor, wherein the secondcrystalline solid is ≥90% (Z)-endoxifen; and (b) optionally, subjectingthe second crystalline solid to recrystallization from ethanol or columnchromatographic treatment one or more times to form a third crystallinesolid.

In some embodiments, the mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III), is prepared by coupling the compound ofFormula (II), (4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, to propiophenone mediated by a McMurry reaction comprisingthe steps of: (a) reacting the compound of Formula (II) withpropiophenone (1:0.01 to 1.5 wt/wt) in an inert organic solvent (1:1 to1:20 wt/wt); (b) preparing a titanium salt (1:0.1 to 1:12 wt/wt) and areducing agent (1:0.01 to 1:10 wt/wt) in an inert organic solvent (1:1to 1:20 wt/wt); and (c) reacting the compound of Formula (II) of step(a) with the titanium salt and a reducing agent in an inert organicsolvent of step (b) to form the mixture of (E)-endoxifen and(Z)-endoxifen; and wherein wt/wt is with respect to the compound ofFormula (II).

In other embodiments, the mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III), is prepared by coupling the compound ofFormula (II), (4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, to propiophenone mediated by a McMurry reaction comprisingthe steps of: (a) reacting the compound of Formula (II) withpropiophenone (1:0.01 to 1:5 wt/wt) in THF (1:1 to 1:20 wt/wt); (b)preparing a TiCl₄ (0.1 to 4 wt/wt) and Zn (0.01 to 1:2 wt./wt) in THF(1:1 to 1:20 wt/wt; and (c) reacting the compound of Formula (II) ofstep (a) with the TiCl₄ and Zn in THF of step (b) to form the mixture of(E)-endoxifen and (Z)-endoxifen; and wherein wt/wt is with respect tothe compound of Formula (II).

In some embodiments, the compound of Formula (II) is generated bydemethylating the compound of Formula (I) with a demethylating agent(1:0.5 to 1:10 wt/wt) and a proton acceptor (1:0.5 to 1:10 wt/wt) in aninert organic solvent (1:1 to 1:20 wt/wt) to form a compound of Formula(II), wherein the wt/wt is with respect to the compound of Formula (I).

In some embodiments, the compound of Formula (II) is prepared bydemethylating the compound of Formula (I) with 1-chloroethylchloroformate and DIPEA in THF to form the compound of Formula (II)comprising the steps of: (a) reacting the compound of Formula (I) withDIPEA (1:0.5 to 1:10 wt/wt) in THF (1:20 wt/wt); (b) adding1-chloroethyl chloroformate (1:0.5 to 1:10 wt./wt); (c) distilling withmethanol one or more times (1:1 to 1:10 wt./wt); (e) reacting withmethanol (1:1 to 1:5 wt/wt)/HCl (1:1 to 1:10 wt/wt); and (e)neutralizing with NaOH (1:1 to 1:10 wt/wt); and wherein wt/wt is withrespect to the compound of Formula (I).

In another aspect, the present disclosure provides process for making(Z)-endoxifen gluconate salts by reacting (Z)-endoxifen with D-gluconateor L-Gluconate to form (Z)-endoxifen L-gluconate or (Z)-endoxifenD-gluconate.

In still another aspect, the present disclosure provides that(Z)-endoxifen, (E)-endoxifen, compounds of Formula (III), the compoundof Formula (II), and salts thereof prepared by any of the processes asdescribed herein are stable. In a particular aspect, the (Z)-endoxifenfree base is stable at ambient temperature for at least 9 months.

In certain aspects, the present disclosure provides an industriallyscalable process for manufacturing crystalline Form II or III of thecompound of Formula (III), comprising (a) reacting to 6N HCL (1:1 to 1:5wt/wt) in ethyl acetate (1:1 to 1:20 wt/wt) a starting mixture of(E)-endoxifen and (Z)-endoxifen having an E/Z-ratio ranging from 99:1 to40:60; (b) neutralizing with 8N NaOH (1:1 to 1:20 wt/wt); (c) washingone or more times with ethyl acetate; (d) washing one or more times witha mixture of ethyl acetate and n-heptane; and (e) recovering crystallineForm II or III of the compound of Formula (III); wherein wt/wt is withrespect the starting mixture of (E)-endoxifen and (Z)-endoxifen.

In yet another aspect, the present disclosure provides a compositioncomprising (Z)-endoxifen free base or a salt thereof prepared accordingto any of the processes described herein. The compositions areformulated for oral, parenteral, topical, and intraductal delivery. Thecompositions are formulated for oral delivery as tablets, caplets,capsules, or pills. In some embodiments, the compositions are formulatedfor oral delivery tablet as enteric tablets, enteric caplets, entericcapsules, delayed-release tablets, delayed-release caplets, anddelayed-release capsules.

In certain aspects, the present disclosure provides compositions havingcertain pharmacokinetic profiles. In one aspect, the present disclosureprovides that at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90% of the endoxifen in the compositions is releasedin intestines. In another aspect, the present disclosure provides thatthe endoxifen in the compositions has a mean half-life of in a subjectranging from 30 hours to 60 hours after administration. In someembodiments, the mean half-life of endoxifen in a subject ranges from 40hours to 55 hours after administration. In another aspect, the presentdisclosure provides that the mean area under the curve extrapolated totime infinity (AUC_(0-inf)) of 200 hr*ng/mL to 10000 hr*ng/mL, of 300hr*ng/mL to 8000 hr*ng/mL, of 400 hr*ng/mL to 6000 hr*ng/mL or of 700hr*ng/mL to 6000 hr*ng/mL.

In another aspect, the present disclosure provides oral compositionscomprising 1 mg to 200 mg per unit dose of (Z)-endoxifen free base or asalt thereof, for administration to a subject in need thereof, whereindaily administration of the oral composition achieves in the subject:(i) a steady state plasma level of endoxifen within 7 to 21 days; or(ii) a steady state plasma level of endoxifen ranging from 25 nM to 300nM; or (iii) a steady state plasma level of endoxifen greater than 30nM; or (iv) maximal plasma levels of endoxifen within 2 to 10 hoursafter administering; or (v) any combination thereof. The presentdisclosure also provides that the compositions are administered to asubject in need thereof for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both in a subject.

In certain aspect, the present disclosure provides methods of treating asubject in need thereof, the method comprising administering to thesubject an oral composition as disclosed herein. In some embodiments,the subject has or is at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both. Invarious embodiments, the hormone-dependent breast disorder or thehormone-dependent reproductive tract disorder is a benign breastdisorder, hyperplasia, atypia, atypical ductal hyperplasia, atypicallobular hyperplasia, increased breast density, gynecomastia, DCIS, LCIS,breast cancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,or vulvar cancer.

In some embodiments, the subject has prostate cancer and wherein thesubject further has or is at risk of having gynecomastia.

In certain aspects, the present disclosure provides certain patientpopulations that would benefit from such methods of treatment. In someembodiments, the patient population is a tamoxifen-refractory ortamoxifen resistant population. In certain embodiments, the subject hastamoxifen-refractory or tamoxifen resistant hormone-dependent breastdisorder or hormone-dependent reproductive tract disorder. In otherembodiments, the patient population comprises subjects that are or willbe treated with an SSRI drug selected from the group consisting ofcitalopram, escitalopram, fluoxetine, paroxetine, sertraline, andvilazodone.

In an aspect, the present disclosure provides certain doses of(Z)-endoxifen to be administered to subjects in need thereof. In variousembodiments, the subjects are administered 0.01 mg to 200 mg of(Z)-endoxifen. In certain embodiments, the subject is administered 1 mg,2 mg, 4 mg, 6 mg, 10 mg or 20 mg of (Z)-endoxifen daily.

In another aspect, the present disclosure provides methods of treatmentof subjects, wherein a steady state plasma level of endoxifen in of thesubject is greater than 30 nM upon administration of the compositions.In some embodiments, such a steady state plasma level of endoxifen isachieved within 7 to 21 days of the first administration of thecomposition. In some embodiments, the time to maximum plasma levels ofendoxifen ranges from 2 hours to 10 hours or from 4 hours to 8 hoursafter administering the composition.

In another aspect, the present disclosure provides methods of treating asubject having or at risk of having a hormone-dependent breast disorder,a hormone-dependent reproductive tract disorder, or both, the methodcomprising administering an oral composition comprising (Z)-endoxifen ora salt thereof, wherein administration of the composition achieves: (i)a mean half-life of endoxifen in the subject ranging from 30 hours to 60hours after administration; (ii) a time to maximum plasma levels ofendoxifen ranging from 4 hours to 8 hours after administration; and(iii) a steady state plasma level of endoxifen greater than 30 nM. Incertain embodiments, the subject is administered 1 mg, 2 mg, 4 mg, 6 mg,10 mg or 20 mg of (Z)-endoxifen. In other embodiments, the mean areaunder the curve extrapolated to time infinity (AUC_(0-inf)) of 200hr*ng/mL to 10000 hr*ng/mL, of 300 hr*ng/mL to 8000 hr*ng/mL, of 400hr*ng/mL to 6000 hr*ng/mL or of 700 hr*ng/mL to 6000 hr*ng/mL. In someembodiments, the composition is formulated as an enteric tablet, anenteric caplet, an enteric capsule, a delayed-release tablet, adelayed-release caplet or a delayed-release capsule. In someembodiments, at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90% of the endoxifen is released in intestines. Insome embodiments, the composition is administered once a day, twice aday, thrice a day, four times a day, every other day, twice a week,weekly, fortnightly, twice a month, monthly, quarterly, once every sixmonths, or annually. In some embodiments, the hormone-dependent breastdisorder and the hormone-dependent reproductive tract disorder areselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer.

INCORPORATION BY REFERENCE

All publications, patents, and patent applications mentioned in thisspecification are herein incorporated by reference to the same extent asif each individual publication, patent, or patent application wasspecifically and individually indicated to be incorporated by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the invention are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present invention will be obtained by reference to thefollowing detailed description that sets forth illustrative embodiments,in which the principles of the invention are utilized, and theaccompanying drawings of which:

FIG. 1 is a scheme depicting the metabolic route of tamoxifen toendoxifen and other metabolites. The plasma concentrations of4-hydroxytamoxifen are low relative to those of N-desmethyl tamoxifen,indicating that the primary route for metabolism of the parent drug isvia N-demethylation by cytochrome P450 3A4 (CYP3A4) to N-desmethyltamoxifen, followed by hydroxylation by cytochrome P450 2D6 (CYP2D6) toendoxifen.

FIG. 2 is a representative infrared spectrum of (Z)-endoxifen free baseprepared using methods disclosed herein.

FIG. 3 shows the time to maximum serum levels (nM) of endoxifenfollowing treatment with placebo or 1 mg, 2 mg or 4 mg endoxifencapsules.

FIG. 4 shows the time to maximum serum levels of endoxifen followingtreatment with placebo or 1 mg, 2 mg or 4 mg endoxifen capsules at day 1and day 21. The time post-dose (hours) is plotted on the X-axis and meanserum concentration (ng/mL) is plotted on the Y-axis.

FIG. 5 shows the time to achieve steady state serum levels of endoxifenfollowing treatment with placebo or 1 mg, 2 mg or 4 mg endoxifencapsules from day 0 (day 0 refers to day 8 on the X-axis) to day 21. TheX-axis is time in days (Day 8 to Day 21) and the Y-axis is mean serumconcentration in ng/mL. Blood was drawn on each day from day 0 to theend of the clinical trial period. Subjects were dosed on day 1 andplasma endoxifen levels were measured as shown in Table 17. Subjectswere subsequently dosed daily starting on day 8 (labeled day 8 in FIG.5) until the end of study period.

FIG. 6 shows mean serum concentrations (ng/mL) over time followingtreatment with placebo or 1 mg, 2 mg or 4 mg endoxifen capsules at day 1and day 21 (Linear).

FIG. 7 shows mean serum concentrations (ng/mL) over time followingtreatment with placebo or 1 mg, 2 mg or 4 mg endoxifen capsules at day 1and day 21 (Semi-log).

FIG. 8 shows mean serum concentrations (nM) over time followingtreatment with placebo or 1 mg, 2 mg or 4 mg endoxifen capsules at day 1and day 21 (Linear).

FIG. 9 is an XRPD pattern obtained from a sample of Form I of thecompound of Formula (III).

FIG. 10 is an XRPD pattern obtained from a sample of Form I of thecompound of Formula (III).

FIG. 11 is an XRPD pattern obtained from a sample of Form II of thecompound of Formula (III).

FIG. 12 is an XRPD pattern obtained from a sample of Form II of thecompound of Formula (III).

FIG. 13 is an XRPD pattern obtained from a sample of Form III of thecompound of Formula (III).

DETAILED DESCRIPTION OF THE INVENTION

Compounds are described using standard nomenclature. Unless otherwisedefined, all technical and scientific terms used herein have the samemeaning as is commonly understood by one of skill in the art to whichthis invention belongs.

As used herein, the terms “a,” “an,” and “the” include plural referenceunless the context dictates otherwise.

As used herein, the terms “active pharmaceutical ingredient”, “activeingredient”, “API,” “drug,” “active,” “actives” or “therapeutic agent”may be used interchangeably to refer to the pharmaceutically activecompound(s) in a pharmaceutical composition. This is in contrast toother ingredients in the compositions, such as excipients, which aresubstantially or completely pharmaceutically inert. A suitable API inaccordance with the present disclosure is one where there is or likelymay be patient compliance issues for treating a certain disease,condition, or disorder. The therapeutic agent as used herein includesthe active compound and its salts, prodrugs, and metabolites. As usedherein the term “drug” means a compound intended for use in diagnosis,cure, mitigation, treatment, and/or prevention of disease in man orother animals.

As used herein, “adjuvant therapy” refers to a therapy that follows aprimary therapy and that is administered to subjects at risk ofrelapsing. Adjuvant systemic therapy in case of breast cancer orreproductive tract cancer, for example with tamoxifen, usually beginssoon after primary therapy to delay recurrence, prolong survival or curea subject.

As used herein, the term “tamoxifen” refers to(Z)-2-[4-(1,2-diphenyl-1-butenyl)phenoxy]-N,N-dimethylethanamine.Tamoxifen can also refer to the E-isomer or a combination of theE-isomer and the Z-isomer.

As used herein, the terms “4-hydroxytamoxifen,” “afimoxifene,” and“4-OHT” used interchangeably refer to4-1-[4-[2-(dimethylamino)ethoxy]phenyl]-2-phenylbut-1-enyl]phenol, andconstitutes an active metabolite of tamoxifen. 4-OHT can refer to theZ-isomer, E-isomer or a combination thereof.

As used herein, the term “endoxifen” refers to4-hydroxy-N-desmethyl-tamoxifen. It is a secondary active metabolite oftamoxifen.

Embodiments that reference throughout this specification to “acompound”, such as compounds of Formula (I), Formula (II), Formula (III)and Formula (IV), include the polymorphic, salt, free base, co-crystal,and solvate forms of the formulas and/or compounds disclosed herein.Thus, the appearances of the phrases “a compound”, “compound of Formula(I)”, “compound of Formula (II)”, “compounds of Formula (III)” and“compound of Formula (IV)” include Form I of the compound of Formula(IV), Forms II-III of the compounds of Formula (III), the free base ofthe compound of Formula (IV), the free base of the compounds of Formula(III), and/or the gluconate salts as described herein.

The terms “crystalline form”, “polymorph” and “Form” may be usedinterchangeably herein, and are meant to include all crystalline andamorphous forms of the compound, including, for example, polymorphs,pseudopolymorphs, salts, solvates, hydrates, unsolvated polymorphs(including anhydrates), conformational polymorphs, and amorphous forms,as well as mixtures thereof, unless a particular crystalline oramorphous form is referred to. Compound of the present disclosureinclude crystalline and amorphous forms of those compounds, including,for example, polymorphs, pseudopolymorphs, salts, solvates, hydrates,unsolvated polymorphs (including anhydrates), conformational polymorphs,and amorphous forms of the compounds, as well as mixtures thereof.

All compounds disclosed herein are further understood to include allpossible isotopes of atoms occurring in the compounds. Isotopes includethose atoms having the same atomic number but different mass numbers. Byway of example, and without limitation, isotopes of hydrogen includetritium and deuterium and isotopes of carbon include ¹¹C, ¹³C and ¹⁴C.

As used herein and in the claims, the terms “comprising,” “containing,”and “including” are inclusive, open-ended and do not exclude additionalunrecited elements, compositional components or method steps.Accordingly, the terms “comprising” and “including” encompass the morerestrictive terms “consisting of” and “consisting essentially of.”

As used herein, the term “combination therapy” refers to the use of acomposition described herein in combination with one or more additionaltreatment. Treatment in combination therapy can be any treatment such asany prophylactic agent, therapeutic agent (such as chemotherapy),radiotherapy, surgery and the like. The combination can refer toinclusion of a therapeutic or prophylactic agent in a same compositionas a composition disclosed herein (for example, in the same capsule,tablet, ointment, etc.) or in separate compositions (for example, in 2separate capsules). The separate compositions may be in a differentdosage form. The use of the terms “combination therapy” and “incombination with” does not restrict the order in which a compositiondescribed herein and prophylactic and/or therapeutic agent and/ortreatment are administered to a subject in need thereof. Compositions ofthe present disclosure can be administered prior to (e.g., 1 minute(min), 5 min, 15 min, 30 min, 45 min, 1 hour (h), 2 h, 4 h, 6 h, 8 h, 10h, 12 h, 24 h, 36 h, 48 h, 72 h, 96 h, 1 week (wk), 2 wk, 3 wk, 4 wk, 5wk, 6 wk, 8 wk, 12 wk, 6 months (m), 9 m, or 1 year before), concomitantwith, or subsequent to (e.g., 1 minute (min), 5 min, 15 min, 30 min, 45min, 1 hour (h), 2 h, 4 h, 6 h, 8 h, 10 h, 12 h, 24 h, 36 h, 48 h, 72 h,96 h, 1 week (wk), 2 wk, 3 wk, 4 wk, 5 wk, 6 wk, 8 wk, 12 wk, 6 months(m), 9 m, or 1 year after) administration of one or more prophylacticand/or therapeutic agent and/or treatment to a subject in thereof.Combination therapy as used herein can also refer to treatment of asubject having a single disease or multiple diseases, for example,prostate cancer in men and gynecomastia.

As used herein, the term “test sample” means sample of blood obtainedfrom a subject. It is to be understood that when blood sample isobtained from a subject, subject's blood is used for determining thesubject's endoxifen levels and/or other biomarkers that may be measuredor tested. As used herein “plasma endoxifen” is used to refer toendoxifen levels in the subject's test sample, whether the test isconducted on whole blood, plasma or serum.

As used herein, the term “dosage form” means the form in which thecompounds or compositions of the present disclosure are delivered to apatient.

As used herein, the term “pharmaceutically acceptable” or“pharmacologically acceptable” means materials, compositions, orvehicles that are compatible with other ingredients of the formulationand that they do not substantially produce adverse reactions, e.g.,toxic, allergic, or immunological reactions, when administered to asubject. They may be approved by a regulatory agency, e.g., of the U.S.Federal or state government or listed in the U.S. pharmacopeia or othergenerally recognized pharmacopeia for use in animals, and moreparticularly in humans.

As used herein, the term “pharmaceutically acceptable carrier” or“carrier” means a pharmaceutically acceptable material, composition, orvehicle, such as a liquid or solid filler, diluent, excipient, solvent,or encapsulating material, involved in carrying or transporting one ormore of the compounds of the present disclosure from one tissue, organ,or portion of the body or across the skin.

As used herein, the term “pharmaceutically acceptable salt” refers toany salt (e.g., obtained by reaction with an acid or a base) of acompound of the present disclosure that is physiologically tolerated ina subject (e.g., a mammal, and/or in vivo, ex-vivo, in vitro cells,tissues, or organs). A “salt” of a compound of the present disclosuremay be derived from inorganic or organic acids and bases. Suitable anionsalts include, arecoline, besylate, bicarbonate, bitartarate,butylbromide, citrate, camysylate, gluconate, glutamate,glycollylarsanilate, hexylresorcinate, hydrabamine, hydrobromide,hydrochloride, hydroxynapthanoate, isethionate, malate, mandelate,mesylate, methylbromide, methylbromide, methylnitrate, methyl sulfate,mucate, napsylate, nitrate, pamaoate (Embonate), pantothenate,phosphate/diphosphate, polygalacuronate, salicylate, stearate, sulfate,tannate, teoclate, fatty acid anions, and triethiodide.

Suitable cations include benzathine, clemizole, chloroprocaine, choline,diethylamine, diethanolamine, ethylenediamine, meglumine, piperazine,procaine, aluminum, barium, bismuth, lithium, magnesium, potassium, andzinc.

For the purposes of this application, the salts of the compounds of thepresent disclosure are contemplated to be pharmaceutically acceptablefor therapeutic uses. However, salts of acids and bases that arenon-pharmaceutically acceptable may also be useful, for example, in thepreparation or purification of a pharmaceutically acceptable compound.

As used herein, the term “pharmaceutical composition” means acombination of the active agent (e.g., an active pharmaceutical compoundor ingredient, API) with a carrier, inert or active (e.g., aphospholipid), making the compositions specially suitable for diagnosticor therapeutic uses in vitro, in vivo, or ex vivo.

As used herein “primary therapy” refers to a first line of treatmentupon initial diagnosis of a hormone-dependent breast disorder, ahormone-dependent reproductive tract disorder, or both in a subject.Exemplary primary therapies may involve surgery, a wide range ofchemotherapies, and radiotherapy.

As used herein, the terms “subject,” “patient,” “participant,” and“individual,” may be used interchangeably herein and refer to a mammalsuch as a human. Mammals also include pet animals such as dogs, cats,laboratory animals, such as rats, mice, and farm animals such as cowsand horses. Unless otherwise specified, a mammal may be of any gender orsex.

As used herein, the term “tamoxifen refractory” refers to a subject thathas been dosed daily with tamoxifen for at least 2 days and has a levelof plasma endoxifen of less than 30 nM (e.g., less than 20 nM, less than25 nM, or less than 30 nM). As used herein, the term “tamoxifenresistance” refers to two classes of resistance: (a) de novo resistance,i.e., non-responsiveness to tamoxifen therapy from the beginning of thetreatment, or (b) acquired resistance, i.e., non-responsiveness totamoxifen therapy after initial responsiveness or tamoxifen-dependentgrowth/stimulated growth while continuing to express estrogen receptors(Minsun Chang. Biomol. Ther. 20(3), 256-267 (2012)). The acquiredresistance to tamoxifen may develop as early as 3 m to 1 year to as lateas 5 to 10 years. As used herein, the term “reference plasma endoxifenlevel” refers to a value of 30 nM.

As used herein, the term “unit dosage form” refers to physicallydiscrete units suitable for unitary dosages for subjects, each unitcontaining a predetermined quantity of active material calculated toproduce the desired therapeutic effect, in association with a suitablepharmaceutical excipient.

It is specifically understood that any numerical value cited hereinincludes all values from the lower value to the upper value, i.e., allpossible combination of numerical values between the lowest value andthe highest value enumerated are to be considered to be expressly statedin this application and the endpoint of all ranges are included withinthe range and independently combinable. For example, if a concentrationrange or beneficial range is stated as 1% to 50%, it is intended thatvalues such as 2% to 40%, 10% to 30%, or 1% to 3% etc., are expresslyenumerated in this specification. It is also to be understood that if aconcentration or dose is stated as a specific value such as 1 mg or 10mg, it is intended that it is intended to include 10% variation. Asanother example, a stated concentration of 20% is intended to includevalues ±10%. Yet another example, if a ratio of 1:10 to 10:1 is stated,then it is intended that ratios such as 1:9 to 9:1, from 1:8 to 8:1,from 1:7 to 7:1, from 1:6 to 6:1, from 1:5 to 5:1, from 1:4 to 4:1, from1:3 to 3:1, from 1:2 to 2:1, from 1:1 to 2:1 or from 2:5 to 3:5 etc. arespecifically intended. There are only some examples of what isspecifically intended. Unless specified otherwise, the values of theconstituents or components of the compositions are expressed in weightpercent of each ingredient in the component.

All methods described herein can be performed in a suitable order unlessotherwise indicated or otherwise clearly contradicted by context. Theuse of any and all examples, or exemplary language (e.g., “such as” and“the like”) is intended merely to illustrate the invention and does notpose a limitation on the scope of the invention unless otherwiseclaimed. No language in the specification should be construed as anyindicating any non-claimed element as essential to practice of theinvention as used herein.

As used herein, the terms “hormone-dependent breast disorder,”“hormone-dependent reproductive tract disorder,” “hormone-dependentbreast and reproductive tract disorder” each and collectively include,without limitation, any breast or reproductive tract (gynecologic)disorder that is related to or is sensitive to high estrogen or normalestrogen levels that need to be reduced, disorders withestrogen-receptor positive (ER+) and/or progesterone-receptor positive(PR+) disorders, for example, breast disorders, endometriosis, uterinefibroids (also called leiomyomas) etc. Reproductive tract disordersinclude endometrial, ovarian, cervical, uterus, vaginal, and vulvarcancers. The terms “estrogen-related disorder” and “estrogen-receptorrelated disorder” may be used interchangeably to refer to the foregoinghormone dependent disorders. The disorders may be presented primarily orsecondarily to an underlying disease, for example, prostate cancer orother disorders such as liver diseases. Hormone-dependent breast andreproductive tract disorder include, for example, McCune-Albrightsyndrome, which is a disorder caused by a mutation in the GNAS geneaffecting bones, skin, and several hormone-producing (endocrine)tissues, often resulting in abnormal scar-like (fibrous) tissue in theirbones, a condition called polyostotic fibrous dysplasia, hyperthyroidismin individuals carrying such mutations, and in girls often resulting inprecocious puberty.

As used herein, “breast disorder” means any aberration or aconstellation of aberrations in the breast. Such aberration may beproliferative, non-proliferative, benign or malignant. Breast disordersinclude benign lesions of the breast (e.g., hyperplasia), increasedbreast density, gynecomastia, mastalgia, and breast cancer. Benignbreast lesions include, but are not limited to, hyperplasia, atypia,ductal hyperplasia, lobular hyperplasia, atypical ductal hyperplasia(ADH), and atypical lobular hyperplasia (ALH). While not cancerous, ADHand ALH may be indicative of a predisposition for breast cancer.

Breast density is a breast disorder identified by visual techniques suchas mammography and reflects increased fibroglandular tissue within thebreast, i.e., overgrowth of stromal and epithelial cells in the breast.Breast density is classified in 4 classes—Class A, B, C and D—based onthe degree of severity of the density. It is an independent risk factorfor breast cancer. At least 23 states in the USA require physicians toinform subjects if they have dense breast(s). There is currently notreatment for dense breasts, although subjects are reminded to makehealthy lifestyle choices and undergo regular mammograms to monitorchanges in breast.

Gynecomastia is a common male breast condition reflecting increasedhyperplasia of the breast tissue, including epithelial hyperplasia, withprevalence of asymptomatic gynecomastia of 60% to 90% in neonates, 50%to 60% in adolescents, and up to 70% in men aged 50 to 69 years(Therapeutics and Clinical Risk Management 2011:7, 145-148). Newborngynecomastia usually resolves itself within 4 weeks of birth and atleast half of adolescent males experience gynecomastia with typicalonset of 13 to 14 years of age (Tanner stage 3 or 4). Gynecomastia hasbeen proposed to be a risk factor for male breast cancer.

Further, gynecomastia often presents itself secondarily to an underlyingdisorder such as prostate cancer, cirrhosis and liver disease, malehypogonadism, hyperthyroidism, renal failure and in patients undergoinghemodialysis, Type I diabetes mellitis, etc. Further, medications, suchas anti-androgen medications or certain anti-psychotics, themselves havebeen reported to cause up to 25% of cases of gynecomastia and can becategorized by their hormone-like action. For example, the most commonside effects attributed to bicalutamide, a nonsteroidal antiandrogenused for treatment of prostate cancer, are gynecomastia and breast pain.

As used herein, “breast cancer” means any malignant tumor of breastcells. Breast cancer may be at any stage of breast cancer, includingstages of a pre-cancer, an early stage cancer, a non-metastatic cancer,a pre-metastatic cancer, a locally advanced cancer, and a metastaticcancer. There are several types of breast cancer. Exemplary breastcancers include, but are not limited to, ductal carcinoma in situ(DCIS), lobular carcinoma in situ (LCIS), invasive (or infiltrating)lobular carcinoma (ILC), invasive (or infiltrating) ductal carcinoma(IDC), microinvasive breast carcinoma (MIC), inflammatory breast cancer,ER-positive (ER+) breast cancer, ER-negative (ER−) breast cancer, HER2+breast cancer, triple negative breast cancer (TNBC), adenoid cystic(adenocystic) carcinoma, low-grade adenosquamatous carcinoma, medullarycarcinoma, mucinous (or colloid) carcinoma, papillary carcinoma, tubularcarcinoma, metaplastic carcinoma, or micropapillary carcinoma. A singlebreast cancer tumor can be a combination of these types or be a mixtureof invasive and in situ cancer.

DCIS is the most common non-invasive breast cancer. It involves the celllining of the breast ducts. In DCIS, the cells have not spread beyondthe walls of the duct into the surrounding breast tissue. About 1 in 5new breast cancer cases will be DCIS. LCIS is a pre-cancerous neoplasia.It may be indicative of a predisposition for invasive cancer. LCIS onlyaccounts for about 15% of the in situ (ductal or lobular) breastcancers.

IDC is the most invasive breast cancer. As the name applies, it is acarcinoma that begins in the breast ducts and then invades thesurrounding fatty tissue. About 8 to 10 invasive breast cancers areinfiltrating ductal carcinomas. IDC is often treated by surgery toexcise the cancerous tissue, and radiation therapy. In addition,chemotherapy combined with immunotherapy (e.g., tamoxifen andtratuzumab) is often used to treat IDC. If the tumor is larger than 4cm, then a radical mastectomy may be performed.

ILC is a cancer that develops in the lobules of the breast and hasinvaded the surrounding tissue. About 1 in 10 invasive breast cancer isan ILC. ILC is treated by surgery to excise the cancerous tissue, andradiation therapy. In addition, chemotherapy and immunotherapycombination (e.g., tamoxifen and tratuzumab) is often used as anadjuvant therapy to treat ILC.

Inflammatory breast cancer accounts for about 1% to 3% of all breastcancers. In inflammatory breast cancer, cancer cells block lymph vesselsin the skin, resulting in the beast turning red and feeling warm. Theaffected breast may become larger or firmer, tender, or itchy.Inflammatory breast cancer is treated with chemotherapy, immunotherapy,radiation therapy and in some cases, surgery.

Estrogen Receptor positive (ER+) breast cancer is characterized by thepresence of estrogen receptors on the surface of the cancerous cells.Growth of ER+ cancer cells is associated with the availability ofestrogen (hormone-dependent or hormone sensitive breast cancer).Approximately, 80% of all breast cancers are ER+ breast cancers.Treatment options for ER+ breast cancer include chemotherapeutic agentsthat block estrogen (e.g., tamoxifen).

The present disclosure relates to methods of manufacturing stablepreparations of (Z)-endoxifen free base, mixtures of (E)-endoxifen and(Z)-endoxifen (E/Z-mix), and salts thereof, and uses thereof. Thepresent disclosure further relates to oral compositions comprising(Z)-endoxifen or salts thereof and methods of treating a subject in needthereof. The present disclosure also provides methods of treating asubject having or at risk of having a hormone-dependent breast disorder,a hormone-dependent reproductive tract (gynecologic) disorder, or bothby administering to the subject an oral composition comprising(Z)-endoxifen, E/Z-mix, or a salt thereof.

In an aspect, the present disclosure relates to oral compositionscomprising endoxifen or salts thereof. Endoxifen comprised incompositions of the present disclosure may be (Z)-endoxifen,(E)-endoxifen or a combination thereof. In one aspect, the presentdisclosure relates to compositions comprising stable (Z)-endoxifen freebase.

In one aspect, the present disclosure provides industrially scalablemethods of making (Z)-endoxifen free base, mixtures of (E)-endoxifen and(Z)-endoxifen (E/Z-mix), and salts thereof. In an aspect, theindustrially scalable methods are synthetic methods of making stable(Z)-endoxifen free base, E/Z-mix, and salts thereof. In another aspect,the present disclosure provides methods of making compositionscomprising stable (Z)-endoxifen free base, E/Z-mix, and salts thereof.

In certain aspects, the present disclosure provides crystalline forms of(Z)-endoxifen free base and crystalline forms of mixtures of(E)-endoxifen and (Z)-endoxifen, as well as pharmaceutical compositionsof endoxifen comprising the crystalline forms described herein.

Synthesis of (Z)-endoxifen Free Base

Several methods for the synthetic preparation of endoxifen are known inthe art. For e.g., methods of synthetic preparations of endoxifen andtheir prodrugs and salts are described in U.S. Pat. No. 9,333,190(Ahmad, Jina Pharmaceuticals); WO 2008/070463 (Ahmad, JinaPharmaceuticals), U.S. Pub. No. 2010/0112041 (Ahmad, JinaPharmaceuticals), WO 2012/050263 (Ahmad, Jina Pharmaceuticals), WO2014/141292 (Desai, Intas Pharmaceuticals), WO 2017/070651 (USA/AlchemLab. Corp.); WO 2009/120999A2 (Kushner), U.S. Pat. No. 8,063,249(Kushner, Olema Pharmaceuticals), U.S. Pat. Nos. 7,531,578 and 8,119,695(Forman and Yu), and WO 2012/050263 (Song, CJ Cheiljedang Corp).

Methods of synthetic preparation of endoxifen have also been publishedin research literature. (Gauthier et al., J. Org. Chem, 61, 3890-3893(1996), Fauq et al., Bioorg Med Chem Lett. 2010 May 15;20(10):3036-3038); Stearns et at, J. Natl. Cancer Inst. Vol 95, No. 23,2003; Johnson et al., Breast Cancer Research and Treatment. 85:151-159,2004; Ogawa, et al. Chem. Pharm. Bull. 39, 911-916, 1991). However,there remains an unmet need for large scale industrial scalablemanufacturing. Stable (Z)-endoxifen free base may be prepared accordanceto Schema 1 as further described below and in Examples 1 to 9.

Demethylation

In one aspect, the present disclosure relates to an industriallyscalable process of making substantially purified (Z)-endoxifen freebase, E/Z-mix, or salts thereof comprising demethylating the compound ofFormula (I),[4-[2-(dimethylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone),(available from AstaTech Pharmaceuticals, Inc., China) to form thecompound of Formula (II). Accordingly, in some embodiments, theindustrial process comprises preparing the compound of Formula (II) bydemethylating[4-[2-(dimethylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone) with ademethylating agent and a proton acceptor in an inert organic solvent toform the compound of Formula (II). The industrially scalable process formaking the compound of Formula (II), (Z)-endoxifen free base, E/Z-mix,or salts thereof comprises demethylating the compound of Formula (I) inamounts ranging from 1 mg to 1000 kg per reaction.

In some embodiments, the compound of Formula (I) can be demethylated byreacting the compound of Formula (I) with a demethylating agent. Thedemethylating agent is any agent suitable for the purpose. Examples ofsuitable demethylating agents include N-iodosuccinamide (NIS), ethylchloroformates (such as 1-chloroethyl chloroformate, dichloroethylchloroformate, trichloroethyl chloroformate, α-Chloroethylchloroformate(ACE-Cl)), vinyl chloroformate (VO-Cl), cynogen bromide (BrCN: vonBraun's reaction), diethyl azodicarboxylate, pyridinium chloride, andthe like.

In some embodiments, the demethylating agent is a chloroformate selectedfrom the group consisting of 1-chloroethyl chloroformate, dichloroethylchloroformate, trichloroethyl chloroformate, α-Chloroethylchloroformate(ACE-Cl), and vinyl chloroformate (VO-Cl). In other embodiments, thedemethylating agent is an ethyl chloroformate selected from the groupconsisting of selected from the group consisting of ethylchloroformates, such as 1-chloroethyl chloroformate, dichloroethylchloroformate, trichloroethyl chloroformate, andα-Chloroethylchloroformate (ACE-Cl). In at least one embodiment, thedemethylating agent is 1-chloroethyl chloroformate.

In some embodiments, a demethylating agent is added to the reactionmixture at a wt/wt ratio of the compound of Formula (I) to demethylatingagent ranging from 1:0.5 to 1:10. In other embodiments, thedemethylating agent is present at ratios (wt/wt) of the compound ofFormula (I) to demethylating agent ranging from 1:2: to 1:5. In at leastone embodiment, the demethylating agent is present at a ratio (wt/wt) ofthe compound of Formula (I) to demethylating agent of 1:2. In at leastone embodiment, demethylating agent is present at a ratio (wt/wt) of thecompound of Formula (I) to demethylating agent of 1:3.3. In otherembodiments, the demethylating agent 1-chloroethyl chloroformate ispresent at a ratio (wt/wt) of the compound of Formula (I) todemethylating agent ranging from 1:0.5 to 1:10.

The demethylation reaction can be carried out in an inert organicsolvent suitable for the demethylation reaction in the presence of aproton acceptor. Such inorganic solvents include dichloromethane,dichloroethane, chloroform, carbon tetrachloride, chlorobenzene, diethylether, 1,4-dioxane, tert-butyl methyl ether (TBME), tetrahydrofuran(THF), N,N-dimethylformamide (DMF), N-methylpyrrolidone (NMP), diglyme,nitromethane, 1,2-dimethoxyethane (DME), pyridine, acetone,acetonitrile, benzene, o-xylene, m-xylene, p-xylene, xylenes, hexanes,cyclohexane, heptane, octane, nonane, and decane, or a combinationthereof. In an aspect, the industrially scalable process comprises oneor more inert organic solvents for demethylation at a (wt/wt) ratio ofthe compound of Formula (I) to inert organic solvent ranging from 1:1 to1:50. In some embodiments, the demethylation solvent is THF at a ratioof the compound of Formula (I) to THF (wt/wt) ranging from 1:1 to 1:50.In other embodiments, the THF is present in ratios (wt/wt) of thecompound of Formula (I) to THF ranging from 1:1 to 1:20.

Proton acceptors suitable for the purpose of the present disclosureinclude, but are not limited to carbonates, such as sodium carbonate andpotassium carbonate, and bicarbonates, such as sodium bicarbonate andpotassium bicarbonate, proton sponge, and ethyldiisopropylamine,(N′,N-diisopropyleamine, “DIPEA”). In an aspect, the industriallyscalable process comprises a proton acceptor added to the reactionmixture at a wt/wt ratio of the compound of Formula (I) to protonacceptor ranging from 1:0.5 to 1:10. In some embodiments, the protonacceptor is present in a ratio (wt/wt) of the compound of Formula (I) toproton acceptor of 1:1:8.

One of skill in the art will be readily able to determine other solventsand proton acceptors known in the art suitable for demethylationreaction of the present disclosure.

Demethylating agents, solvents for demethylation reaction, and thecompound of Formula (I) may be added in any order. Each reagent may beadded to a suitable reactor in a single bolus or in multiple boluses andstirred.

In some embodiments, the compound of Formula (I) is charged to asuitable reactor to which inert organic solvent THF and proton acceptorDIPEA are added for demethylation reaction, and cooled to 0° C. to 20°C., followed by addition of one or more demethylating agents. THF isadded at a wt/wt ratio of 1:1 to 1:20, and proton acceptor DIPEA isadded at a wt/wt ratio of 1:0.5 to 1:10 to the demethylation reactionmixture, where wt/wt is with respect to the compound of Formula (I). Insome embodiments, the reaction mixture is cooled to not more than (NMT)15° C. followed by slow addition of one or more demethylating agents.The reaction may be carried out under inert conditions, such as undernitrogen or argon.

The reaction mixture comprising the compound of Formula (I), one or moredemethylating agents (for example, 1-chloroethyl chloroformate) in oneor more inert organic solvents can be heated at temperature ranging from20° C. to 250° C., such as from 40° C. to 80° C., from 50° C. to 230°C., from 50° C. to 120° C., and from 150° C. to 200° C. In someembodiments, the reaction mixture is heated under reflux. In otherembodiments, the compound of Formula (I) is reacted with demethylatingagent and proton acceptor for not less than (NLT) 5 hours, NLT 8 hours,NLT 12 hours, NLT 24 hours, NLT 36 hours, NLT 48 hour and NLT 72 hours.In at least one embodiment, the reaction is heated under reflux NLT 12hours. The reaction may be held and stored until further use. In someembodiments, the reaction mixture is held not more than (NMT) 24 hoursunder reflux conditions while stirring.

The mixture can be subjected to one or more rounds of distillation underreduced pressure. Distillation may be carried out at NMT 100° C., at NMT95° C., at NMT 90° C., at NMT 85° C., at NMT 80° C., or at NMT 70° C.with solvents suitable for distillation such as ethyl acetate, loweralcohols (non-limiting examples include methanol, ethanol, n-propanol,and isopropanol), benzene, acetone, acetonitrile, toluene,dichloromethane, 1,2-dichloroethane, and chloroform. One of skill in theart will be able to readily determine additional suitable solventsuseful for the purpose.

In at least one embodiment, the solvent used for distillation ismethanol. As a non-limiting example, methanol may be used for solventswap for 2 to 5 rounds of distillation under reduced pressure.

Solvents, such as methanol, may be used for distillation at a (wt/wt)ratio of the compound of Formula (I) to solvent ranging from 1:1 to 1:10per round of distillation.

Then, the mixture can be reacted with a solvent/acid mixture by additionof an acid (wt/wt ratio of the compound of Formula (I) to acid rangingfrom 1:1 to 1:10) to the solvent while stirring and heating underreflux. The acid can be any suitable acid. HCl is an example of acidsuitable for the purpose of the present disclosure. Non-limitingexamples of suitable solvent/acid mixtures include methanol/HCl,ethanol/HCl, propanol/HCl, isopropanol/HCl, methanol/sulfuric acid,methanol/phosphoric acid, ethanol/sulfuric acid, ethanol/phosphoricacid, propanol/sulfuric acid, propanol/phosphoric acid,isopropanol/sulfuric acid, isopropanol/phosphoric acid, methanol/aceticacid, ethanol/acetic acid, propanol/acetic acid, isopropanol/aceticacid, methanol/formic acid, ethanol/formic acid, propanol/formic acid,and isopropanol/formic acid. In at least one embodiment, thesolvent/acid mixture is methanol/6N HCl. As a non-limiting example,methanol in methanol/HCl mixture may be added at a wt/wt ratio of thecompound of Formula (I) to methanol in solvent/acid mixture at 1:3.2while HCl in the methanol/HCl mixture may be at a wt/wt ratio of thecompound of Formula (I) to HCl in the solvent/acid mixture at 1:4. In atleast one embodiment, the methanol/6N HCl mixture is added at a wt/wtratio of the compound of Formula (I) to methanol/6N HCl ranging from 1:1to 1:10, such as 1:1 to 1:5.

Distillation may be carried out for NLT 5 hours, NLT 8 hours, NLT 10hours, NLT 12 hours, NLT 14 hours under reduced pressure. Distillationmay be carried out at a temperature NMT 70° C., NMT 75° C., NMT 80° C.,NMT 85° C., NMT 90° C. and NMT 90° C.

The reaction mixture may be held and stored until further use. In someembodiments, the reaction mixture may be held for NMT 24 hours underreflux conditions while stirring.

The reaction mixture can be neutralized with a neutralizing agent suchas sodium hydroxide (NaOH), ammonium hydroxide, aminomethylpropanol andthe like, and filtered. The resulting wet cake can be washed with waterand an organic solvent such as ethyl acetate (EtOAc) to obtain(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, thecompound of Formula (II).

The neutralizing agent may be at a wt/wt ratio of the compound ofFormula (I) to the neutralizing agent ranging from 1:1 to 1:10. In someembodiments, the neutralizing agent is 8N sodium hydroxide at a ratioranging from 1:1 to 1:10. In other embodiments, the neutralizing agentis 8N sodium hydroxide at a ratio ranging from 1:2 to 1:8.

In another aspect, the industrially scalable process for making(Z)-endoxifen free base, E/Z-mix or salts thereof comprises one or moresteps of washing filtered product (the compound of Formula (II)) withpurified water (1:1 to 1:5 wt/wt) and organic solvents such as ethylacetate (EtOAc) (1:0.5 to 1:10 wt/wt) wherein the wt/wt is with respectto the compound of Formula (I). The wet cake is dried under reducedpressure/vacuum. The temperature for can range from 25° C. to 60° C. Insome embodiments, the drying is carried out at a temperature NMT 50° C.

McMurry Reaction

In another aspect, the present disclosure relates to an industriallyscalable process of making (Z)-endoxifen free base, E/Z-mix, and saltsthereof comprising subjecting the compound of Formula (II) to a McMurryreaction to afford an E/Z-mix (i.e., a mixture of (E)-endoxifen and(Z)-endoxifen free bases), compounds of Formula (III).

The McMurry reaction has been used to prepare tamoxifen (European PatentApplication No. 168175). The present disclosure relates to anindustrially scalable process wherein the compound of Formula (II),(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl) methanone, is coupledto propiophenone mediated by a McMurry reaction via titanium salts suchas chloride salts of titanium (for example, titanium trichloride andtitanium tetrachloride (TiCl₄)) and reducing agents in inert organicsolvents to form an E/Z mix of compounds of Formula (III).

Salts of titanium that are useful for the present disclosure includetitanium halides (such as titanium trichloride (TiCl₃), Titaniumtetrachloride (TiCl₄), titanium iodides, titanium bromides, and titaniumfluorides), titanium (IV) trichloride isopropoxide, and titaniumisopropoxide. In some embodiments, the titanium salt is TiCl₄. Titaniumsalts, such as TiCl₄, are added at a wt/wt ratio of the compound ofFormula (II) to titanium salt ranging from 1:0.1 to 1:12.

Reducing agents include zinc, zirconium, vanadium, niobium, molybdenum,tungsten, aluminum, magnesium, potassium, zinc-copper couple, alkali andalkali earth metals, butylium, lithium, and lithium aluminum hydride. Inat least one embodiment, the reducing agent is zinc. The McMurrysynthesis is conveniently carried out using a reducing agent such aszinc at a wt/wt ratio of the compound of Formula (II) to reducing agentranging from 1:0.1 to 1:10. In some embodiments, the ratio of reducingagent is in excess compared to titanium salts.

The McMurry synthesis can be carried out in one or more inert organicsolvents. Inert organic solvents useful for the McMurry reaction includedichloromethane, dichloroethane, chloroform, carbon tetrachloride,chlorobenzene, diethyl ether, 1,4-dioxane, tert-butyl methyl ether(TBME), tetrahydrofuran (THF), N,N-dimethylformamide (DMF),N-methylpyrrolidone (NMP), diglyme, nitromethane, 1,2-dimethoxyethane(DME), pyridine, acetone, acetonitrile, benzene, o-xylene, m-xylene,p-xylene, xylenes, hexanes, cyclohexane, heptane, octane, nonane, anddecane, or a combination thereof. In some embodiments, the inert organicsolvent is at wt/wt ratio of the compound of Formula (II) to solventranging from 1:1 to 1:50. In other embodiments, the inert organicsolvent is at wt/wt ratios of the compound of Formula (II) to solventranging from 1:1 to 1:20. In some embodiments, the inert organic solventused for the McMurry reaction is THF. THF, in some embodiments, is atwt/wt ratio of the compound of Formula (II) to THE ranging from 1:1 to1:20.

It is advantageous to combine titanium salts and reducing agent in theinert organic solvents to create a pre-mix. Titanium salts are added tothe reducing agent and inert organic solvents at such a rate so as tokeep the internal temperature at NMT 75° C., such as NMT 65° C., NMT 55°C., NMT 45° C., NMT 40° C., NMT 35° C., NMT 30° C., NMT 25° C., NMT 20°C., and NMT 15° C. Accordingly, titanium salts and reducing agent in theinert organic solvents are combined to create a pre-mix. In someembodiments, Zn, TiCl₄ and THF are combined to create Zn/TiCl₄/THFmixture. In at least one embodiment, TiCl₄ is added to Zn and THF andmixed keeping the internal temperature NMT 20° C. for NLT 10 min.

Preparation of titanium salt and a reducing agent in an inert organicsolvent may further comprise heating titanium salt and reducing agent inan inert organic solvent to a temperature ranging from 20° C. to 250°C., such as from 40° C. to 80° C., from 50° C. to 230° C., from 50° C.to 120° C., and from 150° C. to 200° C. In some embodiments, titaniumsalt and reducing agent present in an inert organic solvent are heatedat NLT 60° C. In some embodiments, preparation of titanium salt and areducing agent in an inert organic solvent further comprises heatingtitanium salt and reducing agent in an inert organic solvent underreflux. Titanium salt and reducing agent in inert organic solvent areheated under reflux for NLT 30 min, such as NLT 1 hour, NLT 2 hours, NLT4 hours, NLT 6 hours and NLT 8 hours, under inert conditions such asunder N₂ or argon.

It is also advantageous to pre-mix the compound of Formula (II) withinert organic solvent such as THF and propiophenone and then reactingthe compound of Formula (II) to pre-mixed reducing agent/titaniumsalt/solvent mixture such as a Zn/TiCl₄/THF mixture to form a mixture of(E)-endoxifen and (Z)-endoxifen, compounds of Formula (III).Propiophenone can be added at a wt/wt ratio of the compound of Formula(II) to propiophenone ranging from 1:0.01 to 1:5. Inert organic solventcan be at a wt/wt ratio of the compound of Formula (II) to solventranging from 1:1 to 1:20. The compound of Formula (II) of step (a) isreacted with titanium salt and a reducing agent in an organic solventunder reflux for NLT 0.5 hours, NLT 1 hour, NLT 2 hours, NLT 4 hours,NLT 6 hours, NLT 8 hours, NLT 12 hours, NLT 24 hours, and NLT 48 hours.In at least one embodiment, the compound of Formula (II) is mixed withTHF and propiophenone and reacted to a pre-mixture of Zn, TiCl₄, THF andheat under reflux for NLT 2 hours. In another embodiment, the compoundof Formula (II) is mixed with THF and propiophenone and reacted to theZn/THF/TiCL₄ mixture as described above and heated under reflux for NLT8 hours. The reaction mixture may then be allowed to cool to 0° C. to35° C.

The mixture of (E)-endoxifen and (Z)-endoxifen in the reaction mixturecan be then subjected to extractive purification, distillation andcrystallization to afford a purified mixture of (E)-endoxifen and(Z)-endoxifen. The mixture of (E)-endoxifen and (Z)-endoxifen in thereaction mixture can be subject to extractive purification by extractingwith inert organic solvents, such as THF and MeTHF, or by addition ofsalts, such as potassium carbonate, ammonium chloride, sodium chloride,sodium hydroxide, to the reaction mixture and extraction with an inertorganic solvent, such as THF and MeTHF.

In some embodiments, the reaction mixture is extracted one or more timeswith ammonium chloride, such as 25% ammonium chloride (1:1 to 1:30wt/wt); silica (Celite®) bed (1:0.01 to 1:5 wt/wt); and/or solvent, suchas THF (1:1 to 1:10 wt/wt). In other embodiments, the reaction mixtureis extracted one or more times with potassium carbonate (K₂CO₃), such as40% K₂CO₃ (1:1 to 1:10 wt/wt) and MeTHF (1:1 to 1:10 wt/wt). In someembodiments, the E/Z mixture may be further extracted with NaOH, such as1N NaOH (1:1 to 1:20 wt/wt). In at least one embodiment, NaCl (1:0.1 to1:0.5 wt/wt) may be added to 1N NaOH for the extraction step.

In some embodiments, the reaction mixture may be further extracted oneor more times with THF or MeTHF. In at least one embodiment, thereaction mixture is extracted 3 or more times with MeTHF. The applicantshave found MeTHF to be surprisingly suitable for the step of extractionof a mixture of (E)-endoxifen and (Z)-endoxifen, affording higher yieldsof purified mixtures of (Z)-endoxifen and (E)-endoxifen. In at least oneembodiment, the mixture may be still further extracted with 20% sodiumchloride (1:1 to 1:10 wt/wt).

The reaction mixture can be next subjected to 2 to 5 rounds of solventswap and distillation with a suitable solvent, such as ethyl acetate,IPA, and IPA/PPW (1:1 to 1:10 wt/wt with respect to the compound ofFormula (III)). Distillation may be carried out under reducedpressure/vacuum at temperatures ranging from 30° C. to 90° C. In someembodiments, the distillation may be performed at temperature NMT 30°C., NMT 35° C., NMT 40° C., NMT 45° C., NMT 50° C., NMT 55° C., NMT 60°C., NMT 65° C., NMT 70° C., NMT 75° C., NMT 80° C. and NMT 90° C., andfiltered. The filtered product may be washed with solvents such asEtOAc, IPA, IPA/PPW or n-heptane and then crystallized withcrystallization systems such as EtOAc/n-heptane (1:2 v/v) orIPA/n-heptane (1:2.7 v/v) at a wt/wt ratio of a compound of Formula(III) to EtOAc/n-heptane or IPA/n-heptane ranging from 1:1 to 1:20 anddried, for example, at NMT 60° C., to afford a crystalline solid mixtureof (E)-endoxifen and (Z)-endoxifen free bases, compounds of Formula(III).

In yet another aspect, the present disclosure relates to an industriallyscalable method of manufacturing or reequilibriating an E/Z-mix havingan E/Z ratio of approximately 1:1 (45:55 to 55:45). A suitable reactormay be charged with the compounds of Formula (III) dissolved in an inertorganic solvent, such as ethyl acetate, prepared in a McMurry reactionas described above. The compounds of Formula (III) can have an E/Z ratioof 99:1 to 60:40. The mixture can be concentrated at temperaturesranging from 40° C. to 85° C. In some embodiments, the mixture isconcentrated at temperature NMT 75° C. until the volume reaches 5 vol.The mixture is heated to reflux and then cooled to temperatures rangingfrom 40° C. to 60° C. In some embodiments, the temperature of themixture is cooled to 50±5° C. n-heptane, at a ratio of the compounds ofFormula (III) to n-heptane ranging from 1:1 to 1:20, may be added slowlyto the mixture and the mixture may be then cooled to 0±5° C. The mixturemay be stirred at 0±5° C. for NLT 0.5 hours, such as NLT 1 hour, NLT 2hours, NLT 4 hours, NLT 8 hours, NLT 12 hours or NLT 24 hours. Themixture may be filtered and washed with ethyl acetate/n-Heptane (1:2v/v) at a wt/wt ratio of the compounds of Formula (III) to ethylacetate/n-heptane ranging from 1:1 to 1:10. The wet cake may be driedunder reduced pressure to afford E/Z-endoxifen mixture having anE/Z-ratio of approximately 1:1. Drying may be carried out attemperatures ranging from 30° C. to 70° C. In at some embodiments, thewet cake is dried under reduced pressure or vacuum at NMT 60° C. toafford E/Z-endoxifen mixture having an E/Z-ratio of approximately 1:1(45:55 to 55:45).

In still another aspect, the compounds of Formula (III) may be furtherpurified or enriched or reequilibrated to obtain substantially pure(Z)-endoxifen as described below.

Enrichment Purification of (Z)-endoxifen Free Base

In still another aspect, the present disclosure relates to industriallyscalable methods of manufacturing by enrichment and purification(Z)-endoxifen free base. Industrially scalable enrichment andpurification of (Z)-endoxifen may be carried out as described hereinusing the method of Step 3 of exemplary Schema 1, and as furtherdescribed in Examples 1, 2, 4, and 9. The starting mixture of(E)-endoxifen and (Z)-endoxifen used for fractional crystallization canhave any E/Z ratio, for example, E/Z ratio ranging from 99:1 to 1:10. Insome embodiments, the E/Z-ratio of the starting (E)/(Z)-endoxifenmixtures ranges from 30:70 to 70:30. In some embodiments, the E/Z-ratioof the starting (E)/(Z)-endoxifen mixtures ranges from 99:1 to 1:99. Insome embodiments, the E/Z-ratio of the starting (E)/(Z)-endoxifenmixtures is 51:1, 1:1.8 or 1:5.6.

The E/Z-mix can be the compounds of Formula (III) obtained as describedabove or it can be commercially sourced (for example, fromSigma-Aldrich). A mixture of (E)-endoxifen and (Z)-endoxifen (E/Z-mix)is subjected to fractional crystallization to obtain a first crystallinesolid and a first mother liquor enriched with (Z)-endoxifen free base(Example 1). Fractional crystallization is carried using a first solventwhich is capable of triturating endoxifen and its derivatives such that(Z)-endoxifen tends to remain in filtrate. Suitable first solvents arethose that differentially solubilize the endoxifen isomers, and include,without limitation, ethyl acetate, isopropanol, isopropanol/PPW,acetonitrile, acetonitrile/PPW, and dichloromethane. In someembodiments, the first solvent is ethyl acetate. The first solvent, forexample, ethyl acetate, is added at a wt/wt ratio of the compounds ofFormula (III) to first solvent ranging from 1:1 to 1:20. In someembodiments, the the compounds of Formula (III) are dissolved in firstsolvent, and heated to a temperature ranging from 50° C. to 80° C. andcooled to NMT 35° C.

It has been a surprising discovery that acidification of the mixtureenhances the conversion of (E)-endoxifen to (Z)-endoxifen. Accordingly,in some embodiments, the compounds of formula (III) are pretreated withan acid and then neutralized with a base.

As a non-limiting example, a suitable reactor is charged with thecompounds of Formula (III) to which first solvent is added and cooled to0° C.-5° C. Next, an acid such as HCl or TFA may be added slowly. Insome embodiments, the acid is added to the E/Z-endoxifen mixture at awt/wt ratio of the compounds of Formula (III) to acid ranging from 1:1to 1:5. The reaction mixture may then be heated at temperatures rangingfrom 50° C. to 70° C. while stirring. In some embodiments, the reactionis carried out under reflux. The reaction may be carried out for NLT 4hours, such as NLT 6 hours, NLT 12 hours, NLT 24 hours, and NLT 48hours. The reaction mixture is cooled to 0° C.-5° C. and neutralizedwith a neutralizing agent.

In some embodiments, the neutralizing agent is added to the reactionmixture at a wt/wt ratio of the compounds of Formula (III) toneutralizing agent ranging from 1:1 to 1:5. Suitable neutralizing agentsinclude sodium hydroxide, potassium hydroxide, ammonium hydroxide,aminomethylpropanol, and the like. In some embodiments, the neutralizingagent is 8N sodium hydroxide. The pH of the reaction mixture ispreferably alkaline. In some embodiments, the pH is ≥10, such as ≥11 or≥12.

In some embodiments, (Z)-endoxifen is extracted into the organic layerand collected, and the aqueous phase is washed one or more times withthe first solvent such as ethyl acetate (added at a wt/wt ratio of thecompounds of Formula (III) to first solvent ranging from 1:1 to 1:10).The organic layers are pooled, and washed one or more times with brine(20% NaCl; added at a wt/wt ratio of the compounds of Formula (III) toNaCl ranging from 1:1 to 1:10). The organic layer is treated withactivated carbon and filtered over silicon dioxide (Celite®) bed (addedat a wt/wt ratio of the compounds of Formula (III) to silica rangingfrom 1.01 to 1:0.1). The product is washed again one or more times withfirst solvent (added at a wt/wt ratio of the compounds of Formula (III)to first solvent ranging from 1:1 to 1:10), and distilled. Distillationmay be carried out at temperatures ranging from 50° C. to 80° C. In someembodiments, the temperature may be NMT 75° C.

The first mother liquor is enriched in (Z)-endoxifen as seen in Example1 and Tables 2 and 3. In some embodiments, the first mother liquor isenriched by at least 50% as compared to with the E/Z-ratio of themixture of E-endoxifen and Z-endoxifen. In other embodiments, the firstmother liquor is enriched in (Z)-endoxifen by at least 70% as comparedto the E/Z-ratio of the mixture of E-endoxifen and Z-endoxifen.

The first mother liquor may be subjected to recrystallization byconcentrating the first mother liquor, or swapping out the first solventfrom the first mother liquor one or more times to obtain secondcrystalline solid and a second mother liquor (Table 5).Recrystallization is carried out using a second solvent for swapping.The second solvent is added at a wt/wt ratio of the compounds of Formula(III) to second solvent ranging from 1:1 to 1:10 to create a slurry.Suitable second solvents include IPA, IPA/PPW, acetone, acetone/MTBE,ethanol, EtOAc, EtOAc/n-heptane. In some embodiments, the second solventis IPA. Surprisingly, IPA has been found to triturate (Z)-endoxifen intosolid fraction at levels higher than EtOAc. Accordingly, in someembodiments, when the first solvent is EtOAc, the second solvent is IPAor IPA/PPW. This is useful in directing (Z)-endoxifen first into thefiltrate (first mother liquor) from EtOAc, and then into the solidfraction from IPA or IPA/PPW. In at least one embodiment, the secondsolvent is IPA/PPW (Table 7). In yet another embodiment, the secondsolvent is acetone/MTBE. The second crystalline solid is (Z)-endoxifen,(Z)-4-(1-(4-(2-(methylamino)ethoxy)phenyl)-2-phenylbut-1-enyl)phenol,the compound of Formula (IV). (Molecular weight 373.49; Molecularformula: C₂₅H₂₇NO₂; Melting point 139° C.-143° C. The second crystallinesolid is ≥70% (Z)-endoxifen, such as ≥75% (Z)-endoxifen, ≥80%(Z)-endoxifen or ≥90% (Z)-endoxifen. In some embodiments, the secondcrystalline solid is ≥90% (Z)-endoxifen.

The second crystalline solid may optionally be subjected torecrystallization to obtain a third crystalline solid, (Z)-endoxifen(Table 5). The third crystalline solid may be ≥90% (Z)-endoxifen, suchas ≥91% (Z)-endoxifen, ≥92% (Z)-endoxifen, ≥93% (Z)-endoxifen, ≥94%(Z)-endoxifen, ≥95% (Z)-endoxifen, ≥96% (Z)-endoxifen, ≥97%(Z)-endoxifen, ≥98% (Z)-endoxifen, or ≥99% (Z)-endoxifen. In someembodiments, the third crystalline solid is ≥90% (Z)-endoxifen. In someembodiments, the third crystalline solid is ≥95% (Z)-endoxifen. Thisoptional recrystallization is carried out using a third solvent. Thethird solvent is selected from the group consisting of ethanol,methanol, ethyl acetate, IPA, IPA/PPW, acetone, acetone/MTBE andEtOAc/n-heptane.

In an aspect, the present disclosure relates to pre-heating firstsolvent, second solvent, and third solvent prior to use (Table 2). Insome embodiments, one or more of the first solvent, second solvent, andthird solvent can each be independently preheated to a temperatureranging from 40° C. to 80° C. Fractional crystallization andrecrystallization steps may also include steps of distilling at 60° C.to 80° C. and/or cooling the resulting solution to a temperature rangingfrom 0° C. to 35° C.

It is to be understood that in some embodiments, first, second, andthird crystalline solids as well as the second mother liquor obtained asdescribed above may be further subjected to fractional crystallizationand recrystallization as described above one or more time to obtainpurified (Z)-endoxifen. It is also to be understood that first, second,and/or third crystalline solids obtained may be optionally reprocessedusing column chromatography techniques to obtain more (Z)-endoxifen.

In certain embodiments, the industrially scalable methods describedherein independently comprises additional steps or procedures (e.g., toremove reaction by-products, or to workup, isolate or purify reactionproducts) as detailed in examples herein. In some embodiments, the(Z)-endoxifen free base has <2%, <1%, and <0.5% impurity. In otherembodiments, the compounds of Formula (III) have <2%, <1%, and <0.5%impurity.

A person of skill in the art will recognize several parameters of theforegoing process that may be varied in order to obtain a desirableoutcome. These parameters include for example, the methods and means ofpurification of reaction components and solvents; the order of additionof said reaction components and solvents to the reaction mixture,duration of reaction of said reaction components and solvents; andtemperature and rate of stirring, mixing or agitation of the reactioncomponents and solvents during the reactions.

It was found that the process embodied by the methods herein (alsoincluding the particular process steps) fulfills one or more of thefollowing criteria: better stability, safer, simpler, higher yieldingand more economical when compared with the known processes formanufacturing compounds of Formula (II), (III) and (IV). (Z)-Endoxifenprepared by the processes disclosed below is stable for at least 9months at 5° C. and 25° C. at 60% relative humidity (25° C./60% RH) andfor at least 3 moths at 40° C./75% RH (Example 9). Stability at elevatedtemperatures is indicative of long term stability. Accordingly,(Z)-endoxifen free base prepared by the processes of the presentdisclosure are stable for at least 6 months, such as for at least 9months, at least 12 months, or at least 18 months. Further, the processdescribed herein is considered scalable in multi-kilogram operations,making it suitable for commercial production.

Endoxifen Salts

In still another aspect, the present application provides endoxifensalts and methods of making endoxifen salts. Endoxifen salts known inthe art include hydrochloride (Fauq et al., Bioorg Med Chem Lett. 2010May 15; 20(10):3036-3038) and citrate salts of endoxifen (U.S. Pat. No.9,333,190; U.S. Publication No. 2010/0112041) and are being evaluatedfor oral administration to subjects.

In certain embodiments, the present disclosure provides anion salts ofendoxifen selected from the group consisting of arecoline, besylate,bicarbonate, bitartarate, butylbromide, citrate, camysylate, gluconate,glutamate, glycollylarsanilate, hexylresorcinate, hydrabamine,hydrobromide, hydrochloride, hydroxynapthanoate, isethionate, malate,mandelate, mesylate, methylbromide, methylbromide, methylnitrate,methylsulfate, mucate, napsylate, nitrate, pamaoate (Embonate),pantothenate, phosphate/diphosphate, polygalacuronate, salicylate,stearate, sulfate, tannate, Teoclate, and triethiodide.

In other embodiments, the present disclosure provides cation salts ofendoxifen selected from the group consisting of benzathine, clemizole,chloroprocaine, choline, diethylamine, diethanolamine, ethylenediamine,meglumine, piperazine, procaine, aluminum, barium, bismuth, lithium,magnesium, potassium, and zinc.

The endoxifen salts of the present disclosure may be made using any ofthe compounds of Formulae (I), (II), (III) and (IV), prepared asdescribed above.

Endoxifen Gluconate

In some embodiments, the present disclosure provides a gluconate salt ofendoxifen, or a chemical equivalent thereof. Accordingly, in at leastone embodiment, the endoxifen comprised in a composition of the presentdisclosure is an endoxifen gluconate or a chemical equivalent thereof.Unless specified otherwise, it will be understood that when referring toendoxifen gluconate herein, its chemical equivalents will also beencompassed.

For the purposes of the present disclosure, chemical equivalents ofendoxifen gluconate include all anionic, cationic, and non-ionicreaction complexes between the endoxifen molecule and the gluconatemoiety. Such complexes typically react with the hydroxyl group of theendoxifen molecule. The gluconate moiety includes D-gluconic acid,gluconic acid, glycogenic acid, glycan-A-lactone, and the like. In someembodiments, the gluconate moiety is a pharmaceutically acceptablegluconate salt. Such salts include calcium gluconate, sodium gluconate,and salts of alkali metals and alkaline earth metals such as potassiumgluconate, magnesium gluconate, lithium gluconate, and the like.

Both stereoisomers of gluconate, the D-form and the L-form, are embracedin the present disclosure. In some embodiments, endoxifen gluconatecomprised in the compositions of the present disclosure is selected fromthe group consisting of (Z)-endoxifen D-gluconate, (Z)-endoxifenL-gluconate, (E)-endoxifen D-gluconate, (E)-endoxifen L-gluconate andcombinations thereof. In some embodiments, the pharmaceuticalcomposition comprises (Z)-endoxifen L-gluconate. In certain embodiments,the pharmaceutical composition comprises (Z)-endoxifen D-gluconate. Thecompositions may be present as racemic mixtures, pure stereoisomers(e.g., enantiomers and diastereoisomers), stereo-enriched mixtures andthe like unless otherwise indicated. When a particular stereoisomer isshown or named herein, it will be understood by those skilled in the artthat minor amounts of other stereoisomers may be present in thecompositions of the present disclosure unless otherwise indicated,provided that the utility of the composition as a whole is noteliminated by the presence of such other isomers. Individual isomers maybe obtained by numerous methods that are well known in the art,including chiral chromatography using a suitable chiral stationary phaseor support, or by chemically converting them into diastereoisomers,separating the diastereoisomers by conventional means such aschromatography or recrystallization, then regenerating the originalstereoisomer.

The present disclosure also includes methods of making endoxifengluconate or a chemical equivalent thereof, which includes mixingendoxifen with a gluconate moiety to yield endoxifen gluconate. Methodsof making gluconate salts of therapeutic agents are known in the art(for example, U.S. Publication No. 2002/0127665).

In some embodiments, an endoxifen D-gluconate (such as (Z)-endoxifenD-gluconate) salt may be obtained by mixing an ethanolic slurry of theendoxifen (such as (Z)-endoxifen prepared by the methods describedabove) as the free base with an aqueous solution of D-gluconic acid,obtained by hydrolyzing a 20% w/v solution of D-gluconolactone in waterby heating at 70° C. for 15 to 30 min. In some embodiments, a minimumvolume of ethanol is used and 5 ml of the aqueous D-gluconic acidsolution is added per 1 g of endoxifen free base. Stirring is thencontinued until a clear solution is obtained. The required volume ofthis solution is then added to one or more of other excipients toproduce formulations in which the “active ingredient” is (Z)-endoxifenD-gluconate or (Z)-endoxifen L-gluconate salt. The salt may be purifiedby any of the methods known in the art and disclosed herein.

Purified (Z)-endoxifen free base or a mixture of (E)/(Z)-endoxifen isboth useful for the purpose of preparation of the endoxifen gluconatesalt. A solution of (Z)-endoxifen D-gluconate or (Z)-endoxifenL-gluconate may also be purified from a mixture of (E)/(Z)-endoxifengluconate salt as described herein by fractional crystallization orrecrystallization (Examples 1-3) to obtain a solid purified(Z)-endoxifen salt. The solid salt may be stored at −5° C. under N₂until further use.

It will be understood by those skilled in the art that other endoxifensalts can also be used as starting material with gluconate moiety orgluconate salts to yield endoxifen gluconate or chemical equivalentsthereof.

Endoxifen gluconate or chemical equivalent thereof may be prepared using(Z)-endoxifen prepared as described above or with starting reactantsthat are readily available. For example, crude endoxifen HCl (≥98%) andsodium gluconate are readily available from Sigma-Aldrich, USA.

In some embodiments, endoxifen HCl and sodium gluconate are dissolved ina suitable solvent to yield endoxifen gluconate or a chemical equivalentthereof. In at least one embodiment, the composition comprisingendoxifen gluconate also comprises endoxifen HCl. In other embodiments,an (E)/(Z)-endoxifen mixture (compounds of Formula (III)) and sodiumgluconate are dissolved in a suitable solvent to afford endoxifengluconate or a chemical equivalent thereof.

Suitable solvents for making endoxifen gluconate or chemical equivalentsthereof include, but are not limited to, organic solvents such asalcohols, acetones, DMSO, polyethylene glycol, fatty acids and fattyalcohols and their derivatives, hydroxyl acids, pyrrolidones, urea,vegetable oils, animal oils such as fish oils, essential oils, and thelike or mixtures thereof, and water-miscible solvents such as watermiscible alcohols, dimethylsulfoxide, dimethylformamide, water-miscibleether, for example, tetrahydrofuran, water-miscible nitrile, for exampleacrylonitrile, a water miscible ketone such as acetone or methyl ethylketone, an amide such as dimethylacetamide, propylene glycol, glycerin,polyethylene glycol 400, glycofurol, tetraglycol, and the like, ormixtures thereof.

Water-miscible solvents useful for the preparation of endoxifengluconate are glycerin, ethanol, propanol, isopropanol, propyleneglycol, polyethylene glycols, or mixtures thereof. Additional solventsthat are useful include diglycol monoethyl ether (transcutol); alkeleneglycols, such as dipropylene glycol, propylene glycol, polyethyleneglycols such as PEG 300, 400, 3395, 4450 and the like; dimethylisosorbide; and dehydrated alcohol. In some embodiments, the solvent isa dehydrated alcohol, such as absolute alcohol. In certain embodiments,the amount of solvent is sufficient to dissolve the endoxifen (freebase, salts) and gluconate salts. The concentration of the solvent canalso be adjusted as needed. The reaction can be carried out at roomtemperature and at atmosphere of pressure.

The amount of endoxifen free base or endoxifen salt (such as endoxifenHCl and the like) and gluconate salt (such as and sodium gluconate andthe like) that can be used to make endoxifen gluconate or chemicalequivalent thereof can vary depending on the amount of reactants used.The resulting endoxifen gluconate will have endoxifen:gluconate moietyat 1:1 ratio.

In some embodiments, the amount of endoxifen or endoxifen salt used formaking endoxifen gluconate is from 0.01% to 40% (e.g., from 1% to 10%,or from 3% to 5%) by weight of the total composition (wt/wt). In someembodiments, the gluconate salt used for making endoxifen gluconate isfrom 0.01% to 40% (wt/wt) (e.g., from 1% to 10% (wt/wt), or from 3% to5% (wt/wt)). One of skill in the art will be guided by skill andknowledge in the field and the present disclosure, including withoutlimitation, amounts of reactants which are effective to achieve thedesired yields.

Crystalline Forms

In certain aspects, the present disclosure provides crystalline forms ofendoxifen, including crystalline forms of (Z)-endoxifen free base andcrystalline forms of mixtures of (E)-endoxifen and (Z)-endoxifen. Thepresent disclosure further provides pharmaceutical compositions ofendoxifen comprising the crystalline forms described herein. Acrystalline form of endoxifen may provide the advantage ofbioavailability and stability, suitable for use as an active ingredientin a pharmaceutical composition. Variations in the crystal structure ofa pharmaceutical drug substance or active ingredient may affect thedissolution rate (which may affect bioavailability, etc.),manufacturability (e.g., ease of handling, ability to consistentlyprepare doses of known strength) and stability (e.g., thermal stability,shelf life, etc.) of a pharmaceutical drug product or active ingredient.Such variations may affect the preparation or formulation ofpharmaceutical compositions in different dosage or delivery forms, suchas solid oral dosage forms including tablets and capsules. Compared toother forms such as non-crystalline or amorphous forms, crystallineforms may provide desired or suitable hygroscopicity, particle sizecontrols, dissolution rate, solubility, purity, physical and chemicalstability, manufacturability, yield, and/or process control. Thus,crystalline forms of endoxifen may provide advantages such as: improvingthe manufacturing process of an active agent or the stability orstorability of a drug product form of the compound or an activeingredient, and/or having suitable bioavailability and/or stability asan active agent.

The use of certain solvents and fractional crystallization methods hasbeen found to produce different polymorphic forms of endoxifen,including any one or more of polymorphic Forms I, II and II, which mayexhibit one or more favorable characteristics described above. Theprocesses for the preparation of the polymorphs described herein, andcharacterization of these polymorphs are described in greater detailbelow.

Formula III, Form I

In certain aspects, the present disclosure provides polymorphic Form Iof a compound of Formula (III), wherein at least 90% by weight of thecompound of Formula (III) in the composition is the (Z)-isomer (i.e.,(Z)-endoxifen). In some embodiments, polymorphic Form I exhibits anx-ray powder diffraction (XRPD) pattern substantially as shown in FIG. 9or FIG. 10. In some embodiments, polymorphic Form I has an XRPD patterncomprising at least two, at least three, at least four, at least five,or at least six of the major peaks as the XRPD pattern substantially asshown in FIG. 9 or FIG. 10.

The term “substantially as shown in” when referring, for example, to anXRPD pattern, includes a pattern that is not necessarily identical tothose depicted herein, but that falls within the limits of experimentalerror or deviations when considered by one of ordinary skill in the art.The relative intensities of XRPD peaks can vary, depending upon theparticle size, the sample preparation technique, the sample mountingprocedure and the particular instrument employed. Moreover, instrumentvariation and other factors can affect the two theta (2θ) values.Accordingly, when a specified two theta angle is provided, it is to beunderstood that the specified two theta angle can vary by the specifiedvalue ±0.5°, such as ±0.4°, +0.3°, ±0.2°, or ±0.1°. As used herein,“major peak” refers to an XRPD peak with a relative intensity greaterthan 30%, such as greater than 35%. Relative intensity is calculated asa ratio of the peak intensity of the peak of interest versus the peakintensity of the largest peak in the XRPD pattern.

In certain embodiments, polymorphic Form I is characterized by an x-raypowder diffraction pattern comprising major peaks at 16.8±0.3°,17.1±0.3° and 21.8±0.3° two theta. In some embodiments, polymorphic FormI is characterized by an x-ray powder diffraction pattern comprisingpeaks at 16.0±0.3°, 18.8±0.3° and 26.5±0.3° two theta. In someembodiments, polymorphic Form I is characterized by an x-ray powderdiffraction pattern comprising major peaks at 16.8±0.3°, 17.1±0.3° and21.8±0.3° two theta, and at least one peak selected from 16.0±0.3°,18.8±0.3° and 26.5±0.3° two theta. In some embodiments, polymorphic FormI is characterized by an x-ray powder diffraction pattern comprisingpeaks at 12.3±0.3°, 28.0±0.3° and 29.0±0.3° two theta. In someembodiments, polymorphic Form I is characterized by an x-ray powderdiffraction pattern comprising major peaks at 16.8±0.3°, 17.1±0.3° and21.8±0.3° two theta, and at least one peak selected from 12.3±0.3°,28.0±0.3° and 29.0±0.3° two theta. In some embodiments, polymorphic FormI is characterized by an x-ray powder diffraction pattern comprisingmajor peaks at 16.8±0.3°, 17.1±0.3° and 21.8±0.3° two theta, and atleast one peak selected from 12.3 0.3°, 16.0±0.3°, 18.8 0.3°, 26.5±0.3°,28.0±0.3° and 29.0±0.3° two theta. In some embodiments, polymorphic FormI is characterized by an x-ray powder diffraction pattern comprisingmajor peaks at 16.8±0.3°, 17.1±0.3° and 21.8±0.3° two theta, and peaksat 12.3±0.3°, 16.0±0.3°, 18.8 0.3°, 26.5±0.3°, 28.0±0.3° and 29.0±0.3°two theta.

In certain embodiments, the present disclosure provides a compositioncomprising polymorphic Form I. Greater than 90%, 95% or 99% by weight ofthe compound of Formula (III) in the composition may be polymorphic FormI. In some embodiments, the composition comprises 0.01 mg to 200 mg ofpolymorphic Form I. In some embodiments, the composition comprises about1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of polymorphic Form I.

Formula III, Form II

In certain aspects, the present disclosure provides polymorphic Form IIof a compound of Formula (III), wherein the composition comprises the(E)-isomer and the (Z)-isomer of the compound of Formula (III) (i.e.,(E)-endoxifen and (Z)-endoxifen) in an E/Z ratio between 0.9 and 1.3,such as about 1.1. In some embodiments, polymorphic Form II exhibits anx-ray powder diffraction (XRPD) pattern substantially as shown in FIG.11 or FIG. 12. In some embodiments, polymorphic Form II has an XRPDpattern comprising at least two, at least three, at least four, at leastfive, or at least six of the major peaks as the XRPD patternsubstantially as shown in FIG. 11 or FIG. 12.

In certain embodiments, polymorphic Form II is characterized by an x-raypowder diffraction pattern comprising major peaks at 7.0±0.3°, 11.90.3°, 14.0±0.3° and 18.4±0.3° two theta. In some embodiments,polymorphic Form II is characterized by an x-ray powder diffractionpattern comprising a peak at 22.0±0.3° two theta. In some embodiments,polymorphic Form II is characterized by an x-ray powder diffractionpattern comprising major peaks at 7.0±0.3°, 11.9±0.3°, 14.0±0.3° and18.4±0.3° two theta and a peak at 22.0±0.3° two theta. In someembodiments, polymorphic Form II is characterized by an x-ray powderdiffraction pattern comprising at least one peak selected from 6.6±0.3°,13.3 0.3° and 20.0±0.3° two theta. In some embodiments, polymorphic FormII is characterized by an x-ray powder diffraction pattern comprisingmajor peaks at 7.0±0.3°, 11.9±0.3°, 14.0±0.3° and 18.4±0.3° two theta,and at least one peak selected from 6.6±0.3°, 13.3±0.3° and 20.0±0.3°two theta. In some embodiments, polymorphic Form II is characterized byan x-ray powder diffraction pattern comprising major peaks at 7.0±0.3°,11.9±0.3°, 14.0±0.3° and 18.4±0.3° two theta, and at least one peakselected from 6.6±0.3°, 13.3±0.3°, 20.0 0.3° and 22.0±0.3° two theta. Insome embodiments, polymorphic Form II is characterized by an x-raypowder diffraction pattern comprising major peaks at 7.0±0.3°,11.9±0.3°, 14.0±0.3° and 18.4±0.3° two theta, and peaks at 6.6±0.3°,13.3 0.3°, 20.0±0.3° and 22.0±0.3° two theta.

In certain embodiments, the present disclosure provides a compositioncomprising polymorphic Form II. Greater than 90%, 95% or 99% by weightof the compound of Formula (III) in the composition may be polymorphicForm II. In some embodiments, the composition comprises 0.01 mg to 200mg of polymorphic Form II. In some embodiments, the compositioncomprises about 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of polymorphicForm II.

Formula III, Form III

In certain aspects, the present disclosure provides polymorphic Form IIIof a compound of Formula (III), wherein the composition comprises the(E)-isomer and the (Z)-isomer of the compound of Formula (III) (i.e.,(E)-endoxifen and (Z)-endoxifen) in an E/Z ratio between 0.9 and 1.3,such as about 1.1. In some embodiments, polymorphic Form III exhibits anx-ray powder diffraction (XRPD) pattern substantially as shown in FIG.13. In some embodiments, polymorphic Form III has an XRPD patterncomprising at least two, at least three, at least four, at least five,or at least six of the major peaks as the XRPD pattern substantially asshown in FIG. 13.

In certain embodiments, polymorphic Form III is characterized by anx-ray powder diffraction pattern comprising major peaks at 11.9±0.3°,13.9±0.3°, 17.1±0.3° and 17.7±0.3° two theta. In some embodiments,polymorphic Form III is characterized by an x-ray powder diffractionpattern comprising a peak at 25.3±0.3° two theta. In some embodiments,polymorphic Form III is characterized by an x-ray powder diffractionpattern comprising major peaks at 11.9±0.3°, 13.9±0.3°, 17.1±0.3° and17.7±0.3° two theta and a peak at 25.3±0.3° two theta. In someembodiments, polymorphic Form III is characterized by an x-ray powderdiffraction pattern comprising at least one peak selected from18.2±0.3°, 22.5±0.3° and 26.8±0.3° two theta. In some embodiments,polymorphic Form III is characterized by an x-ray powder diffractionpattern comprising major peaks at 11.9±0.3°, 13.9±0.3°, 17.1±0.3° and17.7±0.3° two theta, and at least one peak selected from 18.2±0.3°,22.5±0.3° and 26.8±0.3° two theta. In some embodiments, polymorphic FormIII is characterized by an x-ray powder diffraction pattern comprisingmajor peaks at 11.9±0.3°, 13.9±0.3°, 17.1±0.3° and 17.7±0.3° two theta,and at least one peak selected from 18.2±0.3°, 22.5±0.3°, 25.3±0.3° and26.8±0.3° two theta. In some embodiments, polymorphic Form III ischaracterized by an x-ray powder diffraction pattern comprising majorpeaks at 11.9±0.3°, 13.9±0.3°, 17.1±0.3° and 17.7±0.3° two theta, andpeaks at 18.2±0.3°, 22.5±0.3°, 25.3±0.3° and 26.8±0.3° two theta.

In certain embodiments, the present disclosure provides a compositioncomprising polymorphic Form III. Greater than 90%, 95% or 99% by weightof the compound of Formula (III) in the composition may be polymorphicForm III. In some embodiments, the composition comprises 0.01 mg to 200mg of polymorphic Form III. In some embodiments, the compositioncomprises about 1 mg, 2 mg, 4 mg, 6 mg, 10 mg or 20 mg of polymorphicForm III.

Endoxifen Free Base Compositions

In one aspect, the present disclosure provides stable (Z)-endoxifen freebase or salts thereof, and compositions comprising (Z)-endoxifen freebase or salts thereof. In some embodiments, the pharmaceuticalcomposition comprises endoxifen predominantly as (Z)-endoxifen freebase.

In certain embodiments, compositions may comprise endoxifen as at least0.1%, at least 0.2%, at least 0.3%, at least 0.4%, at least 0.5%, atleast 1%, at least 5%, at least 10%, at least 20%, at least 25%, atleast 30%, at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90%, at least 91%, at least 92%, at least 93%, atleast 94%, at least 95%, at least 96%, at least 97%, at least 98%, atleast 99%, at least 99.5%, at least 99.99%, or 100% of (Z)-endoxifenfree base wt/wt of total endoxifen in the composition. In at least onecomposition, the composition comprises ≥90% of (Z)-endoxifen free basewt/wt of the total endoxifen in the composition. In another embodiment,the composition comprises ≥95% of (Z)-endoxifen free base wt/wt of thetotal endoxifen in the composition. In yet another embodiment, thecompositions comprise ≥96%, ≥97%, ≥98%, ≥99%, or ≥99.5% of (Z)-endoxifenfree base wt/wt of the total endoxifen in the composition.

In other embodiments, compositions comprising endoxifen comprise 0.01%to 20%, 0.05% to 15%, or 0.1% to 10% of (Z)-endoxifen wt/wt or w/v ofthe composition. In at least one embodiment, the compositions comprisingendoxifen comprise 0.01% to 20% of (Z)-endoxifen wt/wt or w/v of thecomposition. In various other embodiments, the compositions comprisingendoxifen comprise 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%,0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%,2%, 3%, 4%, 5%, 10%, or 20% of (Z)-endoxifen wt/wt of the composition.

In an aspect, the compositions comprising (Z)-endoxifen further comprise(E)-endoxifen. In some embodiments, the endoxifen in the composition hasa ratio of (E)-endoxifen to (Z)-endoxifen (E/Z-ratio) of 1:99; 5:95;10:90, 15:85; 20:80, 25:75; 30:70; 40:70, 45:55; 50:50; 55:45; 60:40;65:45; and 70:30. In other embodiments, compositions comprise endoxifenhaving E/Z-ratio ranging from 10:90 to 70:30. In still otherembodiments, compositions comprise endoxifen having E/Z-ratio rangingfrom 45:55 to 55:45.

Unless specifically referred to by the prefix (Z), (E) or (E/Z),endoxifen used generally without a prefix is used herein to include toany or all endoxifen isoforms.

Endoxifen Salt Compositions

In some embodiments, the present disclosure provides compositionscomprising salts of endoxifen. In some embodiments, the presentdisclosure provides compositions comprising pharmaceutically acceptablesalts of endoxifen. Provided herein in certain embodiments arecompositions comprising 1%, 5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%,80%, 90%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.99% or 100% of endoxifensalt.

In some embodiments, the salt is selected from the group consisting ofarecoline, besylate, bicarbonate, bitartarate, butylbromide, citrate,camysylate, gluconate, glutamate, glycollylarsanilate, hexylresorcinate,hydrabamine, hydrobromide, hydrochloride, hydroxynapthanoate,isethionate, malate, mandelate, mesylate, methylbromide, methylbromide,methylnitrate, methyl sulfate, mucate, napsylate, nitrate, pamaoate(Embonate), pantothenate, phosphate/diphosphate, polygalacuronate,salicylate, stearate, sulfate, tannate, Teoclate, triethiodide,benzathine, clemizole, chloroprocaine, choline, diethylamine,diethanolamine, ethylenediamine, meglumine, piperazine, procaine,aluminum, barium, bismuth, lithium, magnesium, potassium, and zinc. Insome embodiments, the salt is endoxifen gluconate. Endoxifen gluconatecan be selected from the group consisting of (Z)-endoxifen D-gluconate,(E)-endoxifen D-gluconate, (Z)-endoxifen L-gluconate, (E)-endoxifenL-gluconate or a combination thereof.

In some embodiments, a composition comprising endoxifen gluconate iscomprised of 10% to 100% of (Z)-endoxifen D-gluconate on a wt/wt basisof total endoxifen gluconate in the composition. In some embodiments, acomposition comprising endoxifen gluconate is comprised of 10% to 100%of (Z)-endoxifen L-gluconate on a wt/wt basis of total endoxifen in thecomposition.

In other embodiments, a composition comprising endoxifen gluconate iscomprised of 10%, 20%, 30%, 40%, 50%, 60%, 65%, 70%, 75%, 80%, 85%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.75%, 99.99%, or100% of (Z)-endoxifen D-gluconate or (Z)-endoxifen L-gluconate withrespect to total endoxifen gluconate. In some embodiments, thecompositions comprise at least 80%, at least 85%, at least 90%, at least91%, at least 92%, at least 93%, at least 94%, at least 95%, at least96%, at least 97%, at least 98%, at least 99%, at least 99.5%, or atleast 99.99% of (Z)-endoxifen D-gluconate, (Z)-endoxifen L-gluconate ora combination thereof.

Provided herein in some embodiments are compositions comprising(Z)-endoxifen D-gluconate and (E)-endoxifen D-gluconate. (Z)-endoxifenD-gluconate and (E)-endoxifen D-gluconate may be present in thecompositions at ratios ranging from 10:90 to 99:1 wt/wt or v/vrespectively. In some embodiments, the ratio of (Z)-endoxifenD-gluconate to (E)-endoxifen D-gluconate is (wt/wt or v/v) 10:90 to 99:1(e.g., 45:55, 50:50, 60:40, 70:30, 80:20, 90:10; 91:9; 92:8; 93:7; 94:8;95:5, 96:4, 97:3, 98:2, 99:1, 99.5:0.5, or 99.99:0.01) respectively. Incertain embodiments, the ratio of (Z)-endoxifen D-gluconate to(E)-endoxifen D-gluconate (wt/wt or v/v) is 90:10; 91:9; 92:8; 93:7;94:8; 95:5, 96:4, 97:3, 98:2, 99:1, 99.5:0.5, or 99.99:0.01. One ofskill in the art will recognize that other combinations of endoxifengluconate isomers are encompassed in the present disclosure.

In some embodiments, a composition comprising endoxifen gluconatecomprises 0.01%, 0.05%, 0.01%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 1%, 2%, 3%,4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%or 20% endoxifen gluconate (wt/wt) or (w/v) of the composition. In someembodiments, a composition comprising endoxifen gluconate comprises0.01%, 0.05%, 0.01%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 1%, 2%, 3%, 4%, 5%,6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19% or 20%(Z)-endoxifen gluconate (wt/wt) or (w/v) of the composition.

The compounds and compositions of the present disclosure can beadministered to a subject in need thereof by any route known in the art,including without limitation, oral, parenteral, topical, and intraductaldelivery. Accordingly, compositions disclosed herein are formulated tobe compatible with the intended route of administration.

In some embodiments, the compositions comprising endoxifen furthercomprise an excipient. Such an excipient can be compatible with theintended route of administration.

Endoxifen Polymorph Compositions

In one aspect, the present disclosure provides compositions comprisingone or more polymorphic forms, such as Form I, Form II, or Form IIdescribed herein, of endoxifen. In some embodiments, the pharmaceuticalcomposition comprises endoxifen predominantly as polymorph Form I. Insome embodiments, the pharmaceutical composition comprises endoxifenpredominantly as polymorph Form II. In some embodiments, thepharmaceutical composition comprises endoxifen predominantly aspolymorph Form III.

In certain embodiments, a composition comprises endoxifen as at least0.1%, at least 0.2%, at least 0.3%, at least 0.4%, at least 0.5%, atleast 1%, at least 5%, at least 10%, at least 20%, at least 25%, atleast 30%, at least 40%, at least 50%, at least 60%, at least 70%, atleast 80%, at least 90%, at least 91%, at least 92%, at least 93%, atleast 94%, at least 95%, at least 96%, at least 97%, at least 98%, atleast 99%, at least 99.5%, at least 99.99%, or 100% of a singlepolymorphic Form of endoxifen, such as Form I, Form II, or Form III,wt/wt of total endoxifen in the composition. In at least onecomposition, the composition comprises ≥90% of a single polymorphic Formof endoxifen, such as Form I, Form II, or Form III, wt/wt of the totalendoxifen in the composition. In another embodiment, the compositioncomprises ≥95% of a single polymorphic Form of endoxifen, such as FormI, Form II, or Form III, wt/wt of the total endoxifen in thecomposition. In yet another embodiment, the composition comprises ≥96%,≥97%, ≥98%, ≥99%, or ≥99.5% of a single polymorphic Form of endoxifen,such as Form I, Form II, or Form III, wt/wt of the total endoxifen inthe composition. When a particular percentage by weight of endoxifen isa single polymorphic form, the remainder of endoxifen in the compositionis some combination of amorphous endoxifen and/or one or morepolymorphic forms of endoxifen excluding the single polymorphic form.When the polymorphic endoxifen is defined as one particular form ofendoxifen, the remainder is made up of amorphous endoxifen and/or one ormore polymorphic forms other than the particular form specified.Examples of single polymorphic forms include Forms I, II and III ofendoxifen, as well as descriptions of a single polymorphic formcharacterized by one or more properties as described herein.

In other embodiments, a composition comprising endoxifen comprises 0.01%to 20%, 0.05% to 15%, or 0.1% to 10% of a single polymorphic Form ofendoxifen, such as Form I, Form II, or Form III, wt/wt or w/v of thecomposition. In at least one embodiment, the composition comprisingendoxifen comprises 0.01% to 20% of a single polymorphic Form ofendoxifen, such as Form I, Form II, or Form III, wt/wt or w/v of thecomposition. In various other embodiments, the composition comprisingendoxifen comprises 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%,0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%,2%, 3%, 4%, 5%, 10%, or 20% of a single polymorphic Form of endoxifen,such as Form I, Form II, or Form III, wt/wt of the composition. In anaspect, a composition comprising a single polymorphic Form of endoxifen,such as Form I, Form II, or Form III, further comprises a secondpolymorphic Form of endoxifen.

Oral Formulations

In some embodiments, a pharmaceutical composition of the presentdisclosure is formulated for oral delivery. Compositions intended fororal use may be prepared in solid or fluid unit dosage forms. In atleast some embodiments, the compositions are formulated for oraldelivery as tablets, caplets, capsules, pills, powders, troches,elixirs, suspensions, syrups, wafers, chewing gums, dragees, lozenges,and the like.

In some embodiments, the oral dosage forms are solid oral dosage formssuch as tablets, caplets, and capsules. In some embodiments, the capsuleis a hard capsule or a soft capsule. In other embodiments, the capsuleis a gelatin capsule, gelatin-free capsule, a “cap-in-cap” capsule,alginate capsule, hydroxypropylmethyl cellulose (HPMC) capsule, apolyvinyl alcohol (PVA) capsule, a hypromellose capsule, or a starchcapsule.

Excipients

In some embodiments, an oral composition comprising (Z)-endoxifen or asalt thereof further comprises one or more excipients. In someembodiments, an oral composition comprising endoxifen or a polymorphthereof further comprises one or more excipients. Accordingly,compositions designed for oral administration can be made with an inertor active excipient or with an edible carrier as disclosed herein.

In various embodiments, the composition provided herein comprises fromabout 1% to about 99.99%, about 5% to about 95%, about 5% to about 90%,about 10% to about 80%, about 15% to about 70%, about 20% to about 60%,from about 30% to about 95%, from about 50% to about 90%, from about 60%to about 90%, from about 60% to about 80%, or from about 70% to about80% by weight of one or more excipients. In certain embodiments, thecomposition provided herein comprises about 99.99%, about 95%, about90%, about 85%, about 80%, about 75%, about 70%, about 65%, about 60%,about 55%, or about 50% by weight of one or more excipients. In certainembodiments, the composition provided herein comprises about 99.99%,about 99%, about 98%, about 97%, about 96%, about 95%, about 94%, about93%, about 92%, about 91%, about 90%, about 89%, about 88%, about 87%,about 86%, or about 85% by weight of one or more excipients. In certainembodiments, the composition provided herein comprises about 85%, about84%, about 83%, about 82%, about 80%, about 79%, about 78%, about 77%,about 76%, about 75%, about 74%, about 73%, about 72%, about 71%, about70%, about 69%, about 68%, about 67%, about 66%, or about 65% by weightof one or more excipients. In certain embodiments, the compositionprovided herein comprises about 55%, about 54%, about 53%, about 52%,about 51%, about 50%, about 49%, about 48%, about 47%, about 46%, orabout 45% by weight of one or more excipients. In certain embodiments,the composition provided herein comprises about 30%, about 29%, about28%, about 27%, about 26%, about 25%, about 24%, about 23%, about 22%,about 21%, or about 20% by weight of one or more excipients.

Examples of excipients that can be used in the compositions formulatedfor oral administration are provided herein and can include, but are notlimited to, one or more of bulking agents, binders, fillers,disintegrating agents, lubricants, glidants, control release agents,enteric coatings, film-forming agents, plasticizers, colorants,sweetners, flavoring agents and the like, or any combination thereof.

Binders suitable for use in the pharmaceutical compositions providedherein include, but are not limited to, sucrose, starches such as cornstarch, potato starch, or starches such as starch paste, pregelatinizedstarch, and starch 1500, PEG 6000, methocel, walocel HM, Luvitec,Luvicaparolactam, Avicel, SMCC, UNIPURE, gelatin, natural and syntheticgums such as acacia, sodium alginate, alginic acid, other alginates,tragacanth, guar gum, cellulose and its derivatives (e.g., ethylcellulose, cellulose acetate, carboxymethyl cellulose calcium, sodiumcarboxymethyl cellulose), polyvinyl pyrrolidone, methyl cellulose,polyvinyl pyrrolidone, hydroxypropyl methyl cellulose, (e.g., Nos 2208,2906, 2910), microcrystalline cellulose, and mixtures thereof. Suitableforms of microcrystalline cellulose include, but are not limited to, thematerials sold as AVICEL PH 101, AVICEL PH 103 AVICEL RC 581, AVICEL PH105 (available from FMC Corporation, American Viscose Division, AvicelSales, Marcus Hook, Pa.), and mixtures thereof. In some embodiments, thebinder is a mixture of microcrystalline cellulose and sodiumcarboxymethyl cellulose. Suitable anhydrous or low moisture excipientsor additives include AVICEL PH 103 and Starch 1500 LM.

Examples of fillers suitable for use in the pharmaceutical compositionsprovided herein include, but are not limited to, talc, calcium carbonate(e.g., granules or powder), sugars such as dextrose, sucrose, lactose, asalt such as calcium carbonate, calcium phosphate, sodium carbonate,sodium phosphate, starches, microcrystalline cellulose, powderedcellulose, cellulosic bases such as methyl cellulose, carboxymethylcellulose dextrates, kaolin, mannitol, silicic acid, sorbitol, starch,pregelatinized starch, and mixtures thereof.

One or more binder or filler in compositions is typically present infrom about 10% to about 99% (wt/wt) of the composition or the dosageform. In some embodiments, binders and/or fillers in a compositioncomprise about 15% to 99%, about 20% to 60%, about 25% to 55%, about 30%to 50%, about 35% to 60%, about 50% to 99% (wt/wt) of the composition.

Disintegrants can be used in the compositions to provide tablets thatdisintegrate when exposed to an aqueous environment. Tablets thatcontain too much disintegrant may disintegrate in storage, while thosethat contain too little may not disintegrate at a desired rate or underthe desired conditions. Thus, a sufficient amount of disintegrant thatis neither too much nor too little to detrimentally alter the release ofthe active ingredients should be used to form solid oral dosage forms.In some embodiments, the disintegrant is deep in the oral solid dosageform to delay disintegration. The amount of disintegrant used variesbased upon the type of formulation, and is readily discernible to thoseof ordinary skill in the art.

Typical compositions comprise from 0.5% to 15% (wt/wt) of disintegrant.In some embodiments, compositions comprise from 1% to 5% (wt/wt) ofdisintegrant in the composition. In another embodiment, the disintegrantis 1% to 25%, 2% to 20%, 5% to 15%, 8% to 12%, or about 10% (wt/wt) ofthe composition.

Disintegrants that can be used in the pharmaceutical compositionsprovided herein include, but are not limited to, agar, alginic acid,calcium carbonate, microcrystalline cellulose, croscarmellose sodium,crospovidone, polacrilin potassium, sodium starch glycolate, potato ortapioca starch, pre-gelatinized starch, other starches, clays, otheralgins, other celluloses, gums, and mixtures thereof.

Lubricants that can be used in the pharmaceutical compositions providedherein include, but are not limited to, calcium stearate, magnesiumstearate, mineral oil, light mineral oil, glycerin, sorbitol, mannitol,polyethylene glycol, other glycols, stearic acid, sodium lauryl sulfate,talc, hydrogenated vegetable oil (e.g., peanut oil, cottonseed oil,sunflower oil, sesame oil, olive oil, corn oil, and soybean oil), zincstearate, magnesium stearate or potassium stearate, ethyl oleate, ethyllaureate, agar, and mixtures thereof. Additional lubricants include, forexample, a syloid silica gel (AEROSIL 200, manufactured by W.R. GraceCo. of Baltimore, Md.), a coagulated aerosol of synthetic silica(marketed by Degussa Co. of Plano, Tex.), CAB O SIL (a pyrogenic silicondioxide product sold by Cabot Co. of Boston, Mass.), Q7-9120 (DowCorning), and mixtures thereof. If used at all, lubricants are typicallyused in an amount of less than 1% (wt/wt) of the compositions or dosageforms into which they are incorporated. In yet another embodiment, thelubricant is 0.1% to 3%, such as 0.5% to 1% (wt/wt), of the composition.

Plasticizers may be added to control the softness or pliability of oraldosage forms such as shell of a capsule, caplet or a tablet and thus,may improve the mechanical properties of the pH-sensitive materials ofthe coatings on the oral dosage forms. Suitable plasticizers, include,without limitation, petroleum oils (for e.g., a paraffinic process oil,a naphthenic process oil, and an aromatic process oil), squalene,squalane, plant oils, (e.g., olive oil, camelia oil, castor oil, talloil, and a peanut oil), silicon oils, dibasic acid esters, (e.g.,dibutyl phthalate, and dioctyl phthalate), liquid rubbers (e.g.,polybutene and a liquid isoprene rubber), liquid fatty acid esters(e.g., isopropyl myristate ISM), hexyl laurate, diethyl sebacate, anddiisopropyl sebacate, triethyl citrate, triacetin, diethylene glycol,polyethylene glycols, polypropylene glycol, phthalates, sorbitol, glycolsalicylate, crotaminton, and glycerin or mixtures thereof. The amount ofplasticizer may vary depending upon the chemical composition of thepharmaceutical preparation. In one embodiment, the at least oneplasticizer is sorbitol, dimethyl isosorbide, or a glycerol. In anotherembodiment, the plasticizer is 1% to 10%, such as 3% to 5% (wt/wt), ofthe composition.

Examples of glidants include, but are not limited to, colloidal siliconedioxide, cellulose, calcium phosphate, di or tri-basic and the like.

As an example of sweetners or sweetening agents include sucrose,saccharin, dextrose, maltose, sugar substitutes, aspartame, xylitol,mannitol, cyclamate, sucralose, maltitol, sorbitol, acesulfame K and thelike.

Examples of flavoring agents include peppermint, methyl salicylate,peppermint, spearment, methyl salicylate, raspberry, red berry,strawberry, pineapple, orange, cherry and the like.

Compositions formulated for oral delivery as disclosed herein, forexample, tablets, caplets, and capsules, may be coated with one or moreenteric coating agent, control release agent or film forming agent tocontrol or delay disintegration and absorption of the compositionscomprising endoxifen or salts thereof in the gastrointestinal tract andthereby provide a sustained action over a longer period of time.Accordingly, in some embodiments, the tablet can be an enteric tablet,the caplet can be an enteric caplet, or the capsule can be an entericcapsule. The enteric tablets, enteric caplets, or enteric capsules ofthe present disclosure may be prepared by techniques known in the art.

Pharmaceutical preparations disclosed herein may comprise a controlrelease agent. Examples of control release agent suitable for useinclude, without limitation, pH-dependent polymers, acid-insolublepolymers, methyl acrylate-methacrylic acid copolymers, cellulose acetatephthalate (CAP), cellulose acetate succinate, hydroxypropyl methylcellulose phthalate, hydroxypropyl methyl cellulose acetate succinate(hypromellose acetate succinate), polyvinyl acetate phthalate (PVAP),methyl methacrylate-methacrylic acid copolymers, shellac, celluloseacetate trimellitate, sodium alginate, zein, waxes, including syntheticwaxes, microcrystalline waxes, paraffin wax, carnauba wax, and beeswax;polyethoxylated castor oil derivatives, hydrogenated oils, glycerylmono-, di-tribenates, glyceryl monostearate, glyceryl distearate, longchain alcohols, such as stearyl alcohol, cetyl alcohol, and polyethyleneglycol; and mixtures thereof. In some embodiments, a time delay materialsuch as glyceryl monostearate or glyceryl distearate may be used. Inother embodiments, the controlled release reagent is a digestible waxysubstance such as hard paraffin wax.

In some embodiments, compositions may comprise one or more ofpH-dependent polymers such as acid insoluble polymers. The pH-dependentpolymers become increasingly permeable above pH 5.0 but are impermeableat pH below 5.0 whereas acid insoluble polymers become soluble inneutral to weakly alkaline conditions. Such control release polymerstarget upper small intestines and colon. Non-limiting examples ofacid-insoluble polymers include cellulose acetate phthalate, celluloseacetate butyrate, hydroxypropyl methyl cellulose phthalate, algenic acidsalts such as sodium or potassium alginate, shellac, pectin, acrylicacid-methylacrylic acid copolymers (commercially available under thetradename EUDRAGIT® L and EUDRAGIT® S from Rohm America Inc.,Piscataway, N.J. as a powder or a 30% aqueous dispersion; or under thetradename EASTACRYL®, from Eastman Chemical Co., Kingsport, Tenn., as a30% dispersion). Additional examples include EUDRAGIT® L100-55,EUDRAGIT® L30D-55, EUDRAGIT® L100, EUDRAGIT® L100 12,5, EUDRAGIT® S100,EUDRAGIT® S12,5, EUDRAGIT® FS 30D, EUDRAGIT® E100, EUDRAGIT® E 12,5, andEUDRAGIT® PO. In at least one embodiment, the composition comprisesEUDRAGIT® L100-55. EUDRAGIT® RS and RL and EUDRAGIT® NE and NM are alsouseful polymers for the purpose of this disclosure. In some embodiments,the composition comprises EUDRAGIT® L30D 55. In another embodiment, thepreparation comprises EUDRAGIT® FS 30D. One of skill in the art willrecognize that at least some acid insoluble polymers listed herein willalso be biodegradable.

For time delay or delayed-release pharmaceutical preparations of oraldosage forms, glyceryl monostearate, glyceryl distearate, andacid-insoluble polymers, for example polymethacrylate pH-sensitivepolymer-based coatings can be used, (e.g., as coating material, i.e.,enteric coating agents, for enteric coating of capsules, caplets, andtablets). Commercial sources for delayed-release oral dosage forms areavailable, for example DRCaps made of hypromellose (HPMC) from Capsugel,USA. Such delayed-release oral dosage forms forms are acid-resistant andcan resist acidity as seen in stomach for at least 30 min, such as forat least 1 hour, for at least 1.5 hour, or for at least 2 hours. Suchdelayed release oral dosage forms can release at least 40%, at least50%, at least 60%, at least 70%, at least 80% or at least 90% of theendoxifen or salts thereof in the intestines (small intestines, largeintestine/colon etc).

In an aspect of the present disclosure, the enteric tablets, entericcaplets, and enteric capsules may be uncoated. Hard uncoated capsuleswith enteric capability using intrinsically enteric capsule technology(for example, EnTrinsic Drug Delivery available from Capsugel) aresuitable for the purpose of the present disclosure.

In various embodiments, the enteric tablet is a hard tablet made withfree-flowing powder of (Z)-endoxifen or a salt thereof. In variousembodiments, the enteric capsule is a capsule made with free-flowingpowder of (Z)-endoxifen or a salt thereof. In various embodiments, theenteric tablet is a hard tablet made with free-flowing powder ofendoxifen or a polymorph thereof. In various embodiments, the entericcapsule is a capsule made with free-flowing powder of endoxifen or apolymorph thereof.

In some embodiments, the enteric capsule is a non-animal based capsule,such as a hypromellose capsule (for example, commercially availableself-gelling Vcaps, VCaps Plus, VCaps enteric, other enteric capsulesmade using Xcellodose, ENCODE colonic delivery technology, andEnTrinsic™ drug delivery technology from Capsugel). Other technologiesknown in the art and available commercially (for example, Qualicaps,USA, Nutrascience, USA, etc.) for the formulating enteric forms of oralsolid dosage forms can also be utilized. In at least one embodiment, thecapsule is an API-in-capsule, meaning that the (Z)-endoxifen free baseor salts thereof is filled neat into the capsule. In such API-in-capsuleoral dosage forms, the active ingredient, (Z)-endoxifen or salts thereofcan be free flowing powders or micronized powders. When the dosage formis a capsule, in at least one embodiment, the capsule can be a seamlesscapsule or a banded capsule.

Rapid absorption and bioavailability of the anti-cancer therapeuticssuch as endoxifen that can further reduce the cancer growth rate arehighly desirable. In an aspect, the present disclosure provides that thecompositions are formulated for certain pharmacokinetic (PK) properties.

In one aspect, rapid achievement of maximal and steady state plasmalevels of endoxifen is a particular aspect of the present disclosure.The present disclosure provides compositions that achieve a maximalplasma level of endoxifen ranging within 2 to 30 hours, within 3 to 20hours, within 2 to 10 hours or within 4 to 8 hours after administrationof the compositions. Accordingly, in some embodiments, time to maximal(peak) plasma level of endoxifen ranges from 2 to 10 hours afteradministration of the composition. In some embodiments, the time tomaximal plasma level of endoxifen ranges from 4 to 8 hours afteradministration of a composition disclosed herein.

Rapid achievement of steady-state plasma levels of endoxifen is alsohighly desirable, and a composition of the present disclosure mayprovide a plasma level of endoxifen in a subject administered thecomposition comprising a polymorphic form of endoxifen, (Z)-endoxifen ora salt thereof that rapidly achieves steady state. Steady state plasmalevels can be achieved from day 7 to day 21. In some embodiments, thesteady state plasma levels can be achieved by day 7 (FIG. 5) upon dailyadministration of a composition disclosed herein.

In an aspect, the present disclosure provides that circulating endoxifenreleased from a composition disclosed herein can be cleared faster thantamoxifen. Terminal elimination half life of tamoxifen is said to be 5-7days (Jordan C. Steroids. 2007 November; 72(13): 829-842) and peakconcentration time of tamoxifen is approximately 5 hours post-dose.Endoxifen released from a composition disclosed herein can have aterminal elimination half-life ranging from 30 to 60 hours,significantly lower than tamoxifen. In some embodiments, the meanhalf-life ranges from 40 to 53 hours. The mean ratio of AUC_(24h), (Day21)/AUC_(0-inf) (Day1) typically ranges from 0.7 to 1.2 for compositionscomprising 1 mg to 4 mg (Z)-endoxifen, or a salt thereof. Thus,accumulation of endoxifen released from a compositions disclosed hereindoes not significantly vary over continued treatment.

In another aspect, a composition described herein achieves absorption ofendoxifen that is therapeutically effective.

Area under Curve AUC_((0-24 hr)) (“AUC_(24 hr)”) describes the totalexposure of the subject to a drug from time of dosing (0 hr) over a 24hour period. Compositions comprising (Z)-endoxifen or a salt thereoftypically achieve mean (AUC_(24 hr)) of 150 hr*ng/mL to 600 hr*ng/mL onDay 1 of initial (first) dose of a composition comprising 1 mg to 4 mgof (Z)-endoxifen. Compositions comprising (Z)-endoxifen or a saltthereof typically achieve mean AUC_(24 hr) of 400 hr*ng/mL to 2500hr*ng/mL on Day 21 of initial (first) dose of compositions comprising 1mg to 4 mg of (Z)-endoxifen.

AUC_(0-inf) (“AUC_(0-inf)”), a time-averaged concentration of drugcirculating in the body fluid analyzed (normally plasma, blood orserum), describes the total exposure of the subject to a drug. Thepresent disclosure provides that the exposure of a subject to endoxifen(AUC_(0-inf)) can be dose proportional. In some embodiments, AUC_(0-inf)ranges from 200 hr*ng/mL to 10000 hr*ng/mL. In other embodiments, theAUC_(0-inf) ranges from 300 hr*ng/mL to 8000 hr*ng/mL. In certainembodiments, the AUC_(0-inf) ranges from 400 hr*ng/mL to 6000 hr*ng/mLover the dosing range of 1 mg to 4 mg of (Z)-endoxifen (for example, seeTable 17).

Dissolution of the oral dosage forms disclosed herein is tested by thedissolution tests according to the current methods of USP 711. In someembodiments, the oral dosage forms disclosed herein are protected fromthe acidic environment of the stomach and do not dissolve for at least 2hours, at least 3 hours, at least 4 hours, at least 5 hours, 6 hours, atleast 7 hours or at least 8 hours. In at least one embodiment, the oraldosage forms do not release endoxifen for at least 6 hours. In anotherembodiment, the oral dosage forms do not release endoxifen or a saltthereof for at least 2 hours.

In other embodiments, less than 10% of (Z)-endoxifen in a compositioncomprising endoxifen or a salt thereof disclosed herein is released inthe stomach ater 2 hours after administration; or wherein less than 40%of (Z)-endoxifen is released in the stomach after 4 hours ofadministration; or less than 50% of (Z)-endoxifen is released in thestomach after 6 hours of administration, as tested by a method of USP711.

In another embodiment, a composition disclosed herein releases in thestomach less than 10% of (Z)-endoxifen in 2 hours after administration,less than 40% of (Z)-endoxifen in 4 hours after administration; and lessthan 50% of (Z)-endoxifen in 6 hours after administration, as tested bya method of USP 711.

In yet another embodiment, the composition is formulated to release inthe small intestine, such that at least 10% of endoxifen is releasedafter 4 hours after administration; or at least 30% of endoxifen isreleased after 6 hours after administration; or at least 40% ofendoxifen is released after 7 hours after administration; or at least50% of endoxifen is released after 8 hours after administration, astested by a method of USP 711.

In a further embodiment, as determined by a method of USP 711, thecomposition is formulated to release in the colon at least 50% ofendoxifen after 8 hours after administration.

In still further embodiments, the composition is formulated to releasein the colon at least 20% of endoxifen after 4 hours afteradministration; at least 40% of endoxifen after 6 hours afteradministration; at least 60% of endoxifen after 7 hours afteradministration; or at least 80% of endoxifen after 8 hours afteradministration.

An oral dosage form can be of any shape suitable for oraladministration, such as spherical (0.05-5 mL), oval (0.05-7 mL),ellipsoidal, pear (0.3-5 mL), cylindrical, cubic, regular and/orirregular shaped. An oral dosage form may be of any size suitable fororal administration, for example, size 0, size 2, and the like.

One of skill in the art will further recognize that compositionsdisclosed herein may comprise one or more of the excipients known in theart and disclosed herein in any combination appropriate for a desiredformulation or preparation. Additional excipients may generally be foundin Remington's The Science and Practice of Pharmacy, Meade PublishingCo., United States Pharmacopeia/National Formulary. One of skill in theart will be able to select suitable excipients necessary for thepreparation of the formulations and appropriate dosage forms compatiblewith the route of administration based on his or her skill and knowledgein the art and the disclosures made herein. In all cases, the ultimatedosage form should be sterile and stable under the conditions ofmanufacture and storage.

For formulations of the solid dosage compositions disclosed herein, asthe water activity (A_(w)) is less than 0.75, testing Total AerobicPlate Count (TAC) and USP indicator organism is typically not necessary.The publication, “Microbial Bioburden on Oral Solid Dosage Form,” byJose E. Martinez, Pharmaceutical Technology, February 2002, pages 58 to70, is hereby incorporated by reference in its entirety.

Furthermore, since formulations of the compositions disclosed hereinalso have water activity of less than 0.75, then detailed microbialtesting is typically not necessary. TAC is an estimation of the totalviable aerobic microbes present in a sample of raw material, in-processmaterial, or finished product. Samples are analyzed in accordance withthe most current USP 39 <61>, “Microbiological Examination of NonsterileProducts: Microbial Enumeration Tests.”

Acceptable TAC for oral solid dosage forms (OSDFs) are established forthe formulation of the inventive compositions in terms of alert andaction levels, which could be 1000 cfu g/mL, and 10,000 cfu g/mL,respectively. A TAC that is 20,000 cfu g/mL may be consideredunacceptable.

For other formulations, such as liquid or fluid formulations with wateractivity of less than 0.75, Tests for Specified Microorganisms (S.aureus, Ps. aeruginosa, Salmonella, C. albicans, Clostridia, E. coli andBile Tolerant Gram negative bacteria) in compliance with USP GuidelinesChapter 62 may not need to be performed.

Methods of Use

Compounds of Formulae (I), (II), (III), and (IV), endoxifen saltsdisclosed herein, polymorphic forms of endoxifen disclosed herein, andcompositions comprising them may be used in the manufacture ofmedicaments for use in the treatment of a subject in need thereof, forexample, subjects having or at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both.

The compositions of the present disclosure may be used as a primarytherapy, as a part of a neo-adjuvant therapy (to primary therapy), or aspart of adjuvant therapy regimen, where the intention is to ameliorateor cure a subject having or at risk of having a hormone-dependent breastdisorder, hormone-dependent reproductive tract disorder, or both.

In certain embodiments, the disorder is a hormone-dependent breastdisorder. In other embodiments, the disorder is hormone-dependentreproductive tract disorder. In still other embodiments, the subject hasboth a hormone-dependent breast disorder and a hormone-dependentreproductive tract disorder. In some embodiments, the hormone dependentdisorder is a benign breast disorder, hyperplasia, atypia, atypicalductal hyperplasia, atypical lobular hyperplasia, increased breastdensity, gynecomastia, DCIS, LCIS, breast cancer, endometrial cancer,ovarian cancer, uterine cancer, cervical cancer, vaginal cancer orvulvar cancer.

In some embodiments, the breast disorder is increased breast density.For example, the breast disorder is a class B (formerly Class II), classC (formerly class III) or class D (formerly class IV) breast density.

In some embodiments, the hormone-dependent breast disorder orhormone-dependent reproductive tract disorder is precocious puberty. Inother embodiments, the hormone-dependent breast disorder orhormone-dependent reproductive tract disorder is McCune-AlbrightSyndrome.

In some embodiments, the breast disorder is gynecomastia. In someembodiments, gynecomastia is presented secondarily to an underlyingdisease. Accordingly in some embodiments, the subject also hasunderlying disease selected from the group consisting of prostatecancer, cirrhosis and liver disease, male hypogonadism, hyperthyroidism,renal failure and in patients undergoing hemodialysis, or type Idiabetes mellitis. In certain embodiments, the subject has prostatecancer as the underlying disease, wherein the subject has or is at riskof having gynecomastia.

In certain embodiments, the breast cancer is DCIS, LCIS, ILC, IDC, MIC,inflammatory breast cancer, ER-positive (ER+) breast cancer, HER2+breast cancer, adenoid cystic (adenocystic) carcinoma, low-gradeadenosquamatous carcinoma, medullary carcinoma, mucinous (or colloid)carcinoma, papillary carcinoma, tubular carcinoma, metaplasticcarcinoma, or micropapillary carcinoma. In at least one embodiment, asingle breast cancer tumor may be a combination of the foregoing or be amixture of invasive and in situ cancer.

The present disclosure contemplates the use of the compounds andcompositions disclosed herein at various stages in tumor development andprogression, including the treatment of advanced and/or aggressiveneoplasms, i.e., overt disease in a subject that is not amenable to cureby local modalities of treatment such as surgery or radiotherapy,metastatic disease, or locally advanced disease. Accordingly in someembodiments, the breast cancer is a pre-cancer, an early stage cancer, anon-metastatic cancer, a pre-metastatic cancer, or a locally advancedcancer. In at least one embodiment, the breast disorder is metastaticcancer. In some embodiments, the subject further has prostate cancer.

Current choice for therapeutics for such disorders remains tamoxifen,despite serious adverse effects, poor patient compliance and resistanceto the drug due to low plasma endoxifen levels seen subjects. Suchsubjects may have low endoxifen levels upon dosing with tamoxifen forany number of reasons, such as having CYP gene mutations, for example,in CYP2D6, CYP3A4, or CYP2C9, making them unable to metabolize tamoxifento its active metabolite, endoxifen, or low or dysfunctional estrogenreceptor preventing (or decreasing) sufficient tamoxifen uptake, forother reasons yet to be identified. Reported therapeutic levels ofplasma tamoxifen in subjects dosed with 20 mg of oral tamoxifen is ≥30nM ((Lyon et al. Genet Med. 2012 December; 14(12):990-1000).Notwithstanding the mechanism underlying the low plasma endoxifen in asubject, the compositions of the present disclosure are useful for anycondition wherein a subject has low endoxifen or the subject has or isat a risk of having hormone-dependent breast disorder orhormone-dependent reproductive tract disorder. Therefore, thecompositions of the present disclosure can be particularly important inthe treatment of tamoxifen-resistant, hormone-dependent breast disordersor hormone-dependent reproductive tract disorders.

Provided herein in certain embodiments are patient populations for whomthe pharmaceutical compositions are particularly useful. Thecompositions of the present disclosure are also particularly importantin the treatment of tamoxifen-refractory subjects with hormone-dependentbreast disorders or hormone-dependent reproductive tract disorders.Accordingly, in some embodiments, the compositions disclosed herein areuseful for the treatment of tamoxifen refractory or tamoxifen resistantsubjects having or at risk of having hormone-dependent breast disorders,hormone-dependent reproductive tract disorders, or both. In someembodiments, compositions comprising an endoxifen salt, such asendoxifen gluconate, administered to such subject at the doses disclosedherein, will be advantageous.

Further, Donneyong et al. have shown that drug interactions betweentamoxifen and selective serotonin reuptake inhibitors (S SRI) drugs likeProzac and Paxil (paroxetine) exist and are detrimental to breast cancersubjects (Donneyong et al. BMJ 2016; 354:i5014). The SSRI drugs reduceor stop liver metabolism of tamoxifen to endoxifen in subjects on SSRIdrugs. Thus, provided herein in certain embodiments are patientpopulations being treated or to be treated with SSRI drugs that would bebenefitted by treatment with compositions of the present disclosure.

Compositions disclosed herein administered orally maintain the subject'splasma endoxifen at steady state levels greater than 30 nM, for example,at levels ranging from 30 nM to 80 nM or at levels ranging from 30 nM to300 nM. In some embodiments, the plasma steady state endoxifen levelsare maintained at >40 nM. Maintenance of such a plasma endoxifen atsteady state levels greater than 30 nM is advantageous in that thelikelihood of recurrence (relapse) of hormone-dependent breast disordersor hormone-dependent reproductive tract disorders, particularly breastcancer, at plasma endoxifen levels lower than 30 nM is reduced. It isparticularly advantageous for subjects that are poor-metabolizers oftamoxifen (with plasma endoxifen levels lower than 16 nM), intermediatemetabolizers of tamoxifen (with plasma endoxifen levels lower than 27nM) to be dosed with a composition disclosed herein. It alsoadvantageous for subjects being treated or to be treated withantidepressant drugs such as SSRI drugs such as citalopram (Celexa),escitalopram (Lexapro), fluoxetine (Prozac), paroxetine (Paxil, Pexeva),sertraline (Zoloft), vilazodone (Viibryd) and the like, for example, asubject having or likely to have depression.

Whether a subject is tamoxifen-refractory may be determined by dosing asubject with an initial dosage of tamoxifen and determining thesubject's plasma endoxifen steady state level. Plasma endoxifen steadystate levels in a subject dosed with tamoxifen serves as a biomarker forthe tamoxifen-refractory subjects. The plasma endoxifen levels (acuteand/or steady state) may be determined by obtaining from the subject atest sample, which may be blood sample, collected from the subject afterdosing the subject with tamoxifen. Plasma or serum may be obtained fromblood samples for testing the biomarker endoxifen levels. The initialdosage may comprise administering tamoxifen daily for at least 1 day, 2days, 3 days, 15 days, 1 week, 2 weeks, 4 weeks, 1 month, 2 months, 3months, 4 months, 5 months, or 6 months. The subject may also beadministered with a first composition comprising tamoxifen daily for atleast 1 day, 2 days, 3 days, 15 days, 1 week, 2 weeks, 4 weeks, 1 month,2 months, 3 months, 4 months, 5 months, 6 months, 1 year, 2 years, 3years, 4 years, 5 years or 10 years.

A subject's plasma endoxifen steady state level may be determined bymeasuring endoxifen in a test sample. The subject's plasma endoxifensteady state levels are compared to a reference plasma endoxifen level.For the purposes of the present disclosure, the reference plasma levelis 30 nM. If the subject's plasma endoxifen level is determined to belower than 30 nM, then the subject is defined as tamoxifen-refractory.Such a tamoxifen-refractory subject who has or who may be at risk ofhaving a hormone-dependent breast disorder or hormone-dependentreproductive tract disorder is treated by administering to the subjectan oral composition comprising (Z)-endoxifen or a salt thereof disclosedherein, or a polymorphic form of endoxifen disclosed herein. In someembodiments, the composition administered to such a subject comprises(Z)-endoxifen free base. In some embodiments, the compositionadministered to such a subject comprises a polymorphic form, such asForm I, Form II or Form III, of endoxifen. In other embodiments, thecomposition administered to such a subject comprises endoxifen gluconateselected from the group consisting of (Z)-endoxifen D-gluconate,(Z)-endoxifen L-gluconate, (E)-endoxifen D-gluconate, (E)-endoxifenL-gluconate, or a combination thereof. In other embodiments, thecomposition comprising endoxifen is endoxifen HCl or endoxifen citrate.The present disclosure also contemplates that a subject's plasmaendoxifen levels are tracked or monitored periodically or as necessary.If required, a subject who has been administered an initial dosage oftamoxifen may have his or her plasma endoxifen steady state levelsadjusted by administering a composition comprising endoxifen on anongoing basis based on the test results.

In some embodiments, the subject's tamoxifen-refractory status may bedetermined by determining the subject's tamoxifen-metabolites profilewhich is compared with a reference tamoxifen-metabolite profile as seenin control or normal subjects. Subjects with low plasma endoxifen levelsin subject's tamoxifen-metabolite profile as compared to the referencetamoxifen-metabolite profile are administered an oral compositioncomprising endoxifen or a salt thereof. Such compositions may comprisesynthetically prepared endoxifen.

The plasma endoxifen may be measured by any of method known in the art.The levels of plasma endoxifen in test sample may be determined based onsubject's genes, DNA, RNA, protein, tamoxifen-metabolite profile or acombination thereof. The tamoxifen-metabolites profile can include atleast tamoxifen, 4-OHT, N-desmethyltamoxifen, and/or endoxifen. In someembodiments, the level of plasma endoxifen and/or tamoxifen-metaboliteprofile in the test sample is measured by High Performance LiquidChromatography (HPLC), Gas Chromatography Mass Spectrometry (GC-MS),Liquid Chromatography Mass spectrometry (LC-MS), Liquid ChromatographyTandem Mass spectrometry (LC-MS/MS), immunohistochemistry (IHC),polymerase chain reaction (PCR), quantitative PCR (qPCR), and the like.In some embodiments, the tamoxifen-metabolites profile is predictedbased on the subject's genetic composition. In some embodiments, thesubject's CYP genotype includes, without limitation, analysis of CYP2D6,CYP3A4, CYP2C9 genes. In some embodiments, subject's estrogen receptorlevels may be analyzed. In other embodiments, the determination ofplasma endoxifen may be done by a third party laboratory.

Accordingly, provided herein are methods of maintaining in a subject inneed thereof a plasma endoxifen a level greater than 30 nM byadministering to the subject a composition comprising endoxifen or asalt thereof. In some embodiments, the subject's plasma endoxifen levelis maintained at a steady state level greater than 30 nM. In someembodiments, the subject's plasma endoxifen levels are maintained at asteady state level ranging from 30 nM to 300 nM (for example, from 30 nMto 200 nM, or from 30 nm to 80 nM). In some embodiments, the subject'splasma endoxifen levels are maintained at a steady state level >40 nM.

In another aspect, the subjects may have their test samples tested fortheir biomarker profile that may be indicative or monitoring ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both. Such biomarkers are known in the art andinclude, by way of non-limiting examples, biomarkers such as CYP2D6,BRCA-1, BRCA-2, ER, PR, Her2, uPA, PAI, Tf, p53, Ki67, cytokeratins,cancer tumor antigens, and other biomarkers measured by Mammaprint,OncotypeDx, PAM50, EndoxPredict, MammoStrat, and other diagnostic andpredictive tests. A subject with biomarker profile indicating that thesubject has or is at risk of having a hormone-dependent breast disorder,a hormone-dependent reproductive tract disorder, or both can beadministered a composition disclosed herein. In one aspect, the presentdisclosure provides a method of treating a subject having or at risk ofhaving a hormone-dependent breast disorder, a hormone-dependentreproductive tract disorder, or both, comprising determining a subject'stamoxifen-refractory or tamoxifen-resistant status and administering tothe subject a composition described herein.

In some aspects, provided herein are methods of treating atamoxifen-refractory or tamoxifen-resistant subject, the methodcomprising administration to the subject a composition comprisingendoxifen, or a salt or polymorph thereof.

In some embodiments disclosed herein are methods of treating atamoxifen-refractory subject having or at risk for having ahormone-sensitive breast disorder, a hormone-sensitive reproductivetract disorder, or both, the method comprising administration to thesubject an oral composition comprising endoxifen, or a salt thereof,wherein the subject has plasma endoxifen level of less than 30 nM, lessthan 25 nM, less than 20 nM, less than 15 nM, less than 10 nM, less than5 nM or less than 1 nM. In certain embodiments, the compositioncomprising endoxifen salt is endoxifen gluconate, endoxifen HCl, orendoxifen citrate. In other embodiments, an oral solid dosage formcomprising at least 90% (Z)-endoxifen or a salt thereof is administered.In other embodiments, an oral solid dosage form comprising at least 90%of polymorph Form I, Form II, or Form III of endoxifen is administered.

Also provided herein are methods of treating a tamoxifen-refractorysubject, the method comprising: (a) determining or having determinedplasma endoxifen level in a test sample obtained from the subject; (b)comparing or having compared or having determined the level of plasmaendoxifen in the test sample with a reference plasma endoxifen level;(c) determining or having determined a reduced level of plasma endoxifenin the test sample as compared to the reference plasma endoxifen level;and (d) administering a composition comprising endoxifen or a salt orpolymorph thereof to the subject. The administration of a compositioncomprising endoxifen or a salt or polymorph thereof maintains the levelsof plasma endoxifen in the subject at steady state levels greater than30 nM. In some embodiments, the levels of plasma endoxifen in thesubject are maintained at steady state levels ranging from 30 nM to 80nM.

Provided herein are methods of treating a subject having or at risk ofhaving a hormonal dependent breast disorder or a hormonal dependentreproductive tract disorder, the method comprising: (a) administering tothe subject a first composition comprising tamoxifen; (b) determining orhaving determined the level of plasma endoxifen in a test sampleobtained from the subject; (c) determining or having determined reducedlevel of plasma endoxifen in test sample as compared to a referencelevel of plasma endoxifen; and (d) administering an oral compositiondisclosed herein to the subject. The subject may be administered withthe first composition comprising tamoxifen daily for at least 1 day, 2days, 3 days, 15 days, 1 week, 2 weeks, 4 weeks, 1 month, 2 months, 3months, 4 months, 5 months, 6 months, 1 year, 2 years, 3 years, 4 years,5 years or 10 years. In some embodiments, administration of the oralcomposition comprising endoxifen or a salt or polymorph thereofmaintains the subject's plasma endoxifen at levels greater than 30 nM.In other embodiments, administration of the oral composition comprisingendoxifen or a salt or polymorph thereof maintains the subject's plasmaendoxifen at levels ranging from 30 nM to 300 nM (e.g., from 30 nM to200 nM, or from 30 nM to 80 nM). In some embodiments, the subject isadministered an oral composition comprising (Z)-endoxifen D-gluconate,(Z)-endoxifen L-gluconate, (E)-endoxifen D-gluconate, (E)-endoxifenL-gluconate, or a combination thereof. In other embodiments, the oralcomposition comprising endoxifen salt is endoxifen HCl or endoxifencitrate. In some embodiments, the subject is administered an oralcomposition comprising polymorph Form I, Form II or Form III ofendoxifen disclosed herein.

Provided herein are methods of treating a subject with ahormone-dependent breast disorder or hormone-dependent reproductivetract disorder, the method comprising: (a) dosing the subject with afirst composition comprising tamoxifen; (b) determining or havingdetermined the subject's tamoxifen-metabolites profile in a test sampleobtained from the subject; (c) determining a reduced level of subject'splasma endoxifen based on the subject's tamoxifen-metabolites profile tocompared to a level of reference plasma endoxifen in a referencetamoxifen-metabolites profile; and (d) administering an oral compositioncomprising endoxifen or a salt or polymorph thereof to the subject. Incertain embodiments, the composition comprising endoxifen is endoxifengluconate, endoxifen HCl, or endoxifen citrate. In some embodiments, thecomposition comprises polymorph Form I, Form II, or Form III ofendoxifen.

Provided herein are methods for adjusting plasma endoxifen levels in asubject being treated for hormone-dependent breast disorder orhormone-dependent reproductive tract disorder who has one or more CYP2D6or CYP3A4 mutations or has previously administered with initial dosageof tamoxifen, and who has a plasma endoxifen level less than referenceplasma endoxifen level, the method comprising: (a) measuring thesubject's plasma endoxifen level after initial dosage of tamoxifen; (b)comparing the subject's plasma endoxifen levels to the reference plasmaendoxifen level; (c) administering an oral composition comprisingendoxifen or a salt or polymorph thereof to the subject to maintain thesubject's plasma endoxifen level at levels greater than 30 nM. In someembodiments, administration of the oral composition comprising endoxifenor a salt or polymorph thereof maintains the subject's plasma endoxifenat levels ranging from 30 nM to 300 nM (e.g., from 30 nM to 200 nM, orfrom 30 nM to 80 nM). In some embodiments, the subject's plasmaendoxifen level is maintained at a steady state level. The subject maybe administered with initial dosage of tamoxifen daily for at least 1day, 2 days, 3 days, 15 days, 1 week, 2 weeks, 4 weeks, 1 month, 2months, 3 months, 4 months, 5 months, or 6 months.

Provided herein are methods for adjusting the level of plasma endoxifenin a subject being treated for hormone-dependent breast disorder orhormone-dependent reproductive tract disorder who has been previouslyadministered an initial dosage of tamoxifen and who has a level ofplasma endoxifen less than 30 nM, the method comprising: (a) measuringthe level of plasma tamoxifen-metabolite endoxifen of the subject afterthe initial dosage of tamoxifen; (b) comparing the plasma level oftamoxifen-metabolite endoxifen to the reference level for normaltamoxifen-metabolite endoxifen level; (c) administering an adjustingdosage of a composition comprising synthetically prepared endoxifen,wherein the dosage of synthetically prepared endoxifen is sufficient tomaintain the subject's plasma endoxifen at a level greater than 30 nM.In some embodiments, administration of the second composition comprisinga synthetically prepared endoxifen maintains the subject's plasmaendoxifen at levels ranging from 30 nM to 300 nM (e.g., from 30 nM to200 nM, from 30 nM to 80 nM). In some embodiments, the steps (a) to (c)may be repeated until the subject exhibits a desired plasma level ofendoxifen.

In an aspect, the present disclosure contemplates a method of treating asubject having or at risk of having a hormone-dependent breast disorder,a hormone-dependent reproductive tract disorder, or both, the methodcomprising resection of breast tissue of the subject or administeringradiotherapy to the subject and administering an oral compositioncomprising endoxifen or a salt or polymorph thereof disclosed herein. Inanother aspect, the present disclosure contemplates a method of treatinga subject having or at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both, themethod comprising administering an oral composition disclosed hereinprior to resection of breast tissue of the subject or administeringradiotherapy to the subject.

Dosage to be administered to a subject will be usually in a unit dosageform. Examples of ranges for endoxifen in each dosage unit form are from0.01 mg to 200 mg. Dosage shall generally be an effective amount andequivalent, on a molar basis, of the pharmacologically active (Z)-freeform produced by a dosage formulation upon metabolic release of theactive free drug to achieve its desired pharmacological andphysiological effects. In some embodiments, the compositions comprisingendoxifen or an endoxifen salt or polymorph are administered to thesubject at a dose of 0.01 mg to 200.0 mg. In other embodiments, the oralcompositions comprising endoxifen or an endoxifen salt or polymorph areadministered to the subject at a dose of 1 mg to 200.0 mg. In someembodiments, the oral compositions comprising endoxifen or an endoxifensalt or polymorph are administered to the subject at a dose of 0.01 mg,0.05 mg, 0.1 mg, 0.25 mg, 0.5 mg, 0.75 mg, 1.0 mg, 1.5 mg, 2.0 mg, 4.0mg, 6 mg, 8 mg, 10 mg, 20 mg, 40 mg, 50 mg, 100 mg or 200 mg per unitdose. In certain embodiments, the oral compositions comprising at least90% (Z)-endoxifen (wt/wt) of endoxifen are administered at a dose of 1mg, 2.0 mg, 4.0 mg 6 mg, 8 mg, 10 mg, 20 mg, 40 mg, 50, 100 mg or 200 mgper unit dose. In some embodiments, the compositions comprisingendoxifen gluconate are administered at a dose ranging from 0.01 to 20mg. In some embodiments, a composition comprising (Z)-endoxifenD-gluconate is administered at 0.5 mg, 1 mg, 2 mg 4.0 mg, 6 mg, 8 mg, 10mg, 20 mg, 40 mg, 50 mg, 100 mg, and 200 mg per unit dose. In someembodiments, a composition comprising 1 mg of (Z)-endoxifen D-gluconateis administered. In other embodiments, a composition comprising 1 mg of(Z)-endoxifen L-gluconate is administered. In yet other embodiments, acomposition comprising 2 mg of (Z)-endoxifen D-gluconate and(E)-endoxifen D-gluconate is administered. In certain embodiments, anoral composition comprising at least 90% of a polymorph, such aspolymorph Form I, Form II or Form III, of endoxifen (wt/wt) isadministered at a dose of 1 mg, 2.0 mg, 4.0 mg 6 mg, 8 mg, 10 mg, 20 mg,40 mg, 50, 100 mg or 200 mg per unit dose. In some embodiments, acomposition comprising a polymorphic form, such as Form I, Form II orForm III, of endoxifen is administered at a dose ranging from 0.01 to 20mg.

Breast cancer growth rate studies have shown, using mammographicscreening of subjects with breast cancer, that the breast cancer growthrate in the 25^(th) percentiles of women ages 50 to 59 indicate an unmetneed for fast exposure of the subject to therapeutics (Weeden-Fekjaer etal. Breast Cancer Research200810:R41). Rapid absorption andbioavailability of the anti-cancer therapeutics such as endoxifen thatcan further reduce the cancer growth rate is highly desirable.

In one aspect, rapid achievement of maximal and steady state plasmalevels of endoxifen is a particular aspect of the present disclosure.The present disclosure provides that administration of compositionsdisclosed herein to a subject achieves a maximal plasma level ofendoxifen ranging within 2 to 30 hours, within 3 to 20 hours, within 2to 10 hours or within 4 to 8 hours after administration of thecompositions. Accordingly, in some embodiments, time to maximal (peak)plasma level of endoxifen ranges from 2 to 10 hours after administrationof the compositions. In some embodiments, the time to maximal plasmalevel of endoxifen ranges from 4 to 8 hours after administration ofcomposition disclosed herein.

Rapid achievement of steady-state plasma levels of endoxifen is alsohighly desirable. Plasma levels of endoxifen in a subject administeredwith a composition disclosed herein comprising (Z)-endoxifen or a saltor polymorph thereof rapidly achieves steady state. Steady state plasmalevels can be achieved from day 7 to day 21. In some embodiments, thesteady state plasma levels can be achieved by day 7 (FIG. 5) followingdaily administration of a composition disclosed herein.

In an aspect, the present provides that circulating endoxifen releasedfrom compositions disclosed herein can be cleared faster than tamoxifen.Terminal elimination half life of tamoxifen is said to be 5-7 days(Jordan C. Steroids. 2007 November; 72(13): 829-842) and peakconcentration time of tamoxifen is approximately 5 hours post-dose.Endoxifen released from a composition disclosed herein can have aterminal elimination half-life ranging from 30 to 60 hours,significantly lower than tamoxifen. In some embodiments, the meanhalf-life ranges from 40 to 53 hours. Mean Ratio of AUC_(24 h) (Day21)/AUC_(0-inf) (Day1) typically ranges from 0.7 to 1.2 for compositionscomprising 1 mg to 4 mg (Z)-endoxifen or a salt or polymorph thereof.Thus, accumulation of endoxifen released from the compositions disclosedherein do not significantly vary over continued treatment.

In another aspect, the present disclosure provides that administrationof compositions disclosed herein achieve absorption of endoxifen that istherapeutically effective.

Area under Curve AUC_((0-24 h)) (“AUC_(24 hr)”) describes the totalexposure of the subject to a drug from time of dosing (0 hr) over a 24hour period. Compositions comprising (Z)-endoxifen or a salt thereoftypically achieve mean (AUC_(24 hr)) of 150 hr*ng/mL to 600 hr*ng/mL onDay 1 of initial (first) dose of compositions comprising 1 mg to 4 mg of(Z)-endoxifen. Compositions comprising (Z)-endoxifen or salts thereoftypically achieve mean AUC_(24 hr) of 400 hr*ng/mL to 2500 hr*ng/mL onDay 21 of initial (first) dose of compositions comprising 1 mg to 4 mgof (Z)-endoxifen.

AUC_((0-inf)) (“AUC_(0-inf)”), a time-averaged concentration of drugcirculating in the body fluid analyzed (normally plasma, blood orserum), describes the total exposure of the subject to a drug. Thepresent provides that the exposure of subjects to endoxifen(AUC_(0-inf)) can be dose proportional. In some embodiments, AUC_(0-inf)ranges from 200 hr*ng/mL to 10000 hr*ng/mL. In other embodiments, theAUC_(0-inf) ranges from 300 hr*ng/mL to 8000 hr*ng/mL. In certainembodiments, the AUC_(0-inf) ranges from 400 hr*ng/mL to 6000 hr*ng/mLover the dosing range of 1 mg to 4 mg of (Z)-endoxifen (for example, seeTable 17).

A healthcare professional, such as an attending physician, may adjustthe dosing regimen based on the pharmacokinetic profile of thecomposition in the subject.

In one aspect, the compositions of the disclosure can be used alone orin a combination therapy. For example, compositions disclosed herein maybe used in combination with one or more therapeutic agents as part ofprimary therapy, neoadjuvant therapy, or an adjuvant therapy. It is anaspect of the present disclosure that the compositions of the disclosurecan be used in combination with other therapies such as surgery andradiation as neo-adjuvant or adjuvant therapy. Combinations of thecompositions may act to improve the efficacy of the therapeutic agents,and therefore can be used to improve standard cancer therapies. Forexample, when a subject has prostate cancer and is on bicalutamide orenzalutamide therapy for the treatment of prostate cancer, the subjectis likely to develop gynecomastia a result of the therapy. Thecompositions disclosed herein can be administered as a combinationtherapy to the subject having prostate cancer in order to prevent and/ortreat gynecomastia. As another example, a subject with ER+/Her2+positive breast cancer would be on a combination therapy withtrastuzumab or other oncology drugs such as anti-neoplastics orimmunotherapy, and a composition disclosed herein can be used to treatsuch a subject with ER+/Her2+ positive breast cancer. Accordingly, insome embodiments, the compositions further comprise bicalutamide,enzalutamide or anticancer drugs such as trastuzumab, antineoplasticssuch as capecitabine (Xeloda), carboplatin (Paraplatin), cisplatin(Platinol), cyclophosphamide (Neosar), docetaxel (Docefrez, Taxotere),doxorubicin (Adriamycin), pegylated liposomal doxorubicin (Doxil),epirubicin (Ellence), fluorouracil (5-FU, Adrucil), gemcitabine(Gemzar), methotrexate (multiple brand names), paclitaxel (Taxol),protein-bound paclitaxel (Abraxane), vinorelbine (Navelbine), eribulin(Halaven), ixabepilone (Ixempra), and ATP-cassette binding proteininhibitors.

In another aspect, a composition disclosed herein may comprisetherapeutic agents that increase bioavailability of endoxifen in asubject. P-glycoprotein (P-gp, ABCB1) is a highly efficient drug effluxpump expressed in brain, liver, and small intestine, but also in cancercells, that affects pharmacokinetics and confers therapy resistance formany anticancer drugs. Accordingly, in some embodiments, thecompositions further comprise inhibitors of ATP-binding cassette (ABCfamily) transporters, such as inhibitors of breast cancer resistanceprotein (BCRP protein) and P-gp. Several inhibitors of BCRP protein andP-Gp are known in the art. For example, inhibitors of BCRP proteininclude cyclosporine, omeprazole, pantoprazole, saquinavir, andtacrolimus.

Non-limiting examples of P-gp inhibitors include first generationinhibitors such as Verapamil, cyclosporin A, reserpine, quinidine,yohimbine, tamoxifen and toremifene, second generation inhibitors suchas Dexverapamil, dexniguldipine, valspodar (PSC 833), and Dofequidarfumarate (MS-209), third generation P-gp inhibitors such asCyclopropyldibenzosuberane zosuquidar (LY335979), laniquidar (R101933),mitotane (NSC-38721), biricodar (VX-710), elacridar (GF120918/GG918),ONT-093, tariquidar (XR9576), and HM30181 and anti-P-gp monoclonalantibodies such as MRK-16).

The present disclosure additionally provides for therapeutic kitscontaining one or more of the compositions for use in the treatment of asubject having or at risk of having a hormone-dependent breast disorder,a hormone-dependent reproductive tract disorder, or both. The kits ofthe present disclosure may include an oral composition disclosed herein,a sealed container for housing the composition, and instructions for useof the orally administered composition. In an aspect, the kits of thepresent disclosure can include a second therapeutic agent. Such a secondtherapeutic agent may be bicalutamide, enzalutamide or an anticancerdrug such as trastuzumab, antineoplasitcs such as capecitabine (Xeloda),carboplatin (Paraplatin), cisplatin (Platinol), cyclophosphamide(Neosar), docetaxel (Docefrez, Taxotere), doxorubicin (Adriamycin),pegylated liposomal doxorubicin (Doxil), epirubicin (Ellence),fluorouracil (5-FU, Adrucil), gemcitabine (Gemzar), methotrexate(multiple brand names), paclitaxel (Taxol), protein-bound paclitaxel(Abraxane), vinorelbine (Navelbine), eribulin (Halaven), ixabepilone(Ixempra), and ATP-binding cassette (ABC transporter) inhibitors such asP-gp inhibitors.

Exemplary Compositions

Exemplary, non-limiting compositions are provided below. As mentionedabove, percentages (%) refer to amounts by weight based upon the totalweight of the composition (wt/wt). The sum of the different componentsof the composition adds up to 100% (wt/wt) of the total composition. Theat least 90% (≥90%) (Z)-endoxifen free base refers to the percent weightof (Z)-endoxifen isomer as compared to the total weight of endoxifen inany composition.

In an aspect, the present disclosure relates to industrially scalableprocesses for manufacturing (Z)-endoxifen, compounds of Formula (III),the compound of Formula (II), and salts and polymorphs thereof.

In an aspect, the present disclosure relates to an industrially scalableprocess for manufacturing (Z)-endoxifen, comprising the steps of: (a)reacting a mixture of (E)-endoxifen and (Z)-endoxifen, compounds ofFormula (III), to 6N HCL (1:1 to 1:5 wt/wt) in EtOAc (1:1 to 1:20wt/wt), (b) neutralizing with 8N NaOH (1:1 to 1:20 wt/wt); (c) washingone or more times with EtOAc (1:1 to 1:10 wt/wt); (d) extracting with20% NaCl (1:1 to 1:5 wt/wt); (e) reacting with activated carbon (1:0.01to 1:1); (f) washing one or more times with IPA (1:1 to 1:10 wt/wt) andextracting with MeOH/PPW (1:1 to 1:10 wt/wt); wherein the wt/wt is withrespect to the mixture of (E)-endoxifen and (Z)-endoxifen, e.g.,compounds of Formula (III).

In an aspect, the present disclosure relates to an industrially scalableprocess for manufacturing a mixture of (E)-endoxifen and (Z)-endoxifen(compounds of Formula (III)), comprising: (a) reacting the compound ofFormula (II) with propiophenone in THF (4.4 wt/wt); (b) preparing asolution of TiCl₄ (1.4 wt/wt) and Zn (0.9 wt/wt) in THF (8.9 wt/wt); and(c) reacting the compound of Formula (II) of step (a) with TiCl₄ and Znin THF from step (b) to form a mixture of (E)-endoxifen and(Z)-endoxifen; wherein wt/wt is with respect to the compound of Formula(II).

In an aspect, the present disclosure relates to an industrially scalableprocess for manufacturing a mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III), comprising one or more steps of: (a)extraction with 25% ammonium chloride (1:20 wt/wt) and silica (Celite)(1:1 wt/wt); (b) one or more washes with THF (1:1 to 1:5 wt/wt); (c) oneor more washes with 20% sodium chloride (1:3 wt/wt); (c) distillationwith EtOAc (1:4.5 wt/wt); and (d) crystallization with (1:2 v/v)EtOAc/n-heptane (1:3.8 wt/wt); wherein the wt/wt is with respect to thecompound of Formula (II).

In an aspect, the present disclosure relates to an industrially scalableprocess for manufacturing a mixture of (E)-endoxifen and (Z)-endoxifen,compounds of Formula (III), further comprising one or more steps of: (a)extraction one or more times with 40% K₂CO₃ (1:2 wt/wt); (b) extractionwith 1N NaOH (1:10 wt/wt) and MeTHF (1:1 to 1:10 wt/wt); (c) two moreextractions with MeTHF (1:1 to 1:20 wt/wt); (d) extraction with 20%sodium chloride (1:5 wt/wt); (e) distillation with IPA (1:4.5 wt/wt);and (f) crystallization with (1:2.7 v/v) IPA/n-heptane (1:3.4 wt/wt);wherein the wt/wt is with respect to the compound of Formula (II).

In an aspect, the present disclosure relates to an industrially scalableprocess for manufacturing the compound of Formula (II), comprising thesteps of: (a) reacting the compound of Formula (I) (1 equiv.) with DIPEA(3 wt/wt) in THF (4.9 wt/wt); (b) adding 1-chloroethyl chloroformate(3.3 wt/wt); (c) distilling one to five times with methanol (4.0 wt/wt);(d) distilling with methanol (3.2 wt/wt); (e) reacting with methanol(3.2 wt/wt)/6N HCl (4 wt/wt); and (f) neutralizing with 8N NaOH (5wt/wt); and wherein wt/wt is with respect to the compound of Formula(I).

In an aspect, the present disclosure relates to (Z)-endoxifen,(E)-endoxifen, compounds of Formula (III), and the compound of Formula(II), and salts thereof, prepared according to processes describedherein. In an aspect, the present disclosure relates to (Z)-endoxifenprepared according to a process disclosed herein, wherein the(Z)-endoxifen is stable at ambient temperature for at least 6 months, atleast 9 months, at least 12 months, or at least 18 months.

In another aspect, the (Z)-endoxifen free based prepared in accordancewith the processes disclosed herein have <1% impurity. In still anotheraspect, the (E)/(Z)-endoxifen free base prepared in accordance with theprocesses disclosed herein has <1% impurity.

In an aspect, the present disclosure relates to compositions comprising(Z)-endoxifen free base or a salt thereof prepared according to any ofthe methods disclosed herein.

In another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen free base or a salt thereof prepared accordingto any of the methods disclosed herein, wherein the (Z)-endoxifen is atleast 90% (Z)-endoxifen free base wt/wt of total endoxifen in thecomposition.

In still another aspect, the compositions further comprise(E)-endoxifen, wherein the ratio of (E)-endoxifen to (Z)-endoxifen(E/Z-ratio) is 1:99; 5:95; 10:90, 15:85; 20:80, 25:75; 30:70; 40:70,45:55; 50:50; 55:45; 60:40; 65:45; or 70:30.

In a further aspect, the compositions further comprise (E)-endoxifen,wherein the E/Z-ratio ranges from 45:55 to 55:45 or is approximately1:1.

In an aspect, the compositions comprise (Z)-endoxifen or a salt orpolymorph thereof prepared according to any of the methods disclosedherein, wherein the (Z)-endoxifen is stable at ambient temperature forat least 6 months, at least 9 months, at least 12 months or at least 18months;

In another aspect, the compositions comprise (Z)-endoxifen or saltsthereof prepared according to any of the methods disclosed herein,

-   -   a. wherein the compositions have aerobic bacterial plate count        of not more than 20,000 g/mL;    -   b. wherein water content of the compositions is not more than        1.0% as tested by Method Ic of the USP 921;    -   c. wherein the water activity (Aw) of the compositions is less        than 0.9,    -   d. wherein the residue on ignition is not more than 0.1% as        tested by a method of USP 281;    -   e. wherein a heavy metal is not more than 20 ppm as tested by        Method II of USP 231; or    -   f. wherein methanol is NMT 3000 ppm, tetrahydrofuran is NMT 720        ppm, isopropanol is NMT 5000 ppm, ethyl acetate is NMT 5000 ppm;        n-Heptane is NMT 5000 ppm, and ethanol is NMT 5000 ppm as tested        by a validated HPLC method

In still another aspect, the compositions comprising (Z)-endoxifen orsalts thereof prepared according to any of the methods disclosed hereincomprise 0.01 mg to 200 mg (Z)-endoxifen or a salt or polymorph thereofper unit dose.

In yet another aspect, the compositions comprising (Z)-endoxifen orsalts thereof prepared according to any of the methods disclosed hereincomprise 1 mg to 20 mg of (Z)-endoxifen or a salt or polymorph thereofper unit dose.

In a further aspect, the compositions comprising (Z)-endoxifen or saltsthereof prepared according to any of the methods disclosed hereincomprise 0.01% to 20% (wt/wt) of endoxifen or a salt or polymorphthereof.

In a still further aspect, the present disclosure relates tocompositions comprising (Z)-endoxifen or salts thereof preparedaccording to any of the methods disclosed herein, wherein thecomposition comprises 0.1% to 10% (wt/wt) of (Z)-endoxifen or a salt orpolymorph thereof.

In another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein further comprising one or more excipient.

In yet another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, further comprising one or more excipient,wherein the excipient is a binder, a filler, a disintegrating agent, alubricant, a glidant, a control release agent, an enteric coating agent,a film forming agent, a plasticizer, a sweetening agent, a flavoringagent, or a combination thereof.

In still another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, further comprising one or more excipient,wherein the excipients are about 0.1% to about 99% wt/wt of thecomposition.

In a further aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition comprises one ormore control release agents selected from the group consisting ofacid-insoluble polymers, methyl acrylate-methacrylic acid copolymers,cellulose acetate phthalate (CAP), cellulose acetate succinate,hydroxypropyl methyl cellulose phthalate, hydroxypropyl methyl celluloseacetate succinate (hypromellose acetate succinate), polyvinyl acetatephthalate (PVAP), methyl methacrylate-methacrylic acid copolymers,shellac, cellulose acetate trimellitate, sodium alginate, zein, waxes,including synthetic waxes, microcrystalline waxes, paraffin wax,carnauba wax, and beeswax, polyethoxylated castor oil derivatives,hydrogenated oils, glyceryl mono-, di-tribenates, glyceryl monostearate,glyceryl distearate, long chain alcohols, such as stearyl alcohol, cetylalcohol, polyethylene glycol, and mixtures thereof.

In one aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal, parenteral, topical, and intraductal delivery.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition formulated fororal delivery is a tablet, a caplet, a capsule, a pill, a powder, atroche, an elixir, a suspension, a syrup, a wafer, a chewing gum, adragee, and a lozenge.

In yet another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the compositions formulated fororal delivery are tablets formulated as enteric tablets, capletsformulated as enteric caplets, and capsule formulated as entericcapsules.

In yet another aspect, the present disclosure relates to compositionscomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the compositions formulated fororal delivery are tablets formulated as delayed-release tablets, capletsformulated as delayed-release caplets, or capsules formulated asdelayed-release capsules.

In yet another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in stomach:

-   -   a. less than 10% of endoxifen in 2 hours after administration;    -   b. less than 20% of endoxifen in 2 hours after administration;    -   c. less than 30% of endoxifen in 2 hours after administration;    -   d. less than 40% of endoxifen in 4 hours after administration;        or    -   e. less than 50% of endoxifen in 6 hours after administration.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in stomach:

-   -   a. less than 10% of (Z) endoxifen in 2 hours after        administration;    -   b. less than 20% of (Z) endoxifen in 2 hours after        administration;    -   c. less than 40% of (Z) endoxifen in 4 hours after        administration; or    -   d. less than 50% of (Z) endoxifen in 6 hours after        administration.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases at least 40%, at least 50%, at least 60%, at least70%, at least 80%, or at least 90% endoxifen in the intestines.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in the intestines at least 40%, at least 50%, atleast 60%, at least 70%, at least 80%, or at least 90% endoxifen in 2hours after administration.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in the intestines at least 40%, at least 50%, atleast 60%, at least 70%, at least 80%, or at least 90% endoxifen in 3hours after administration.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in the intestines at least 40%, at least 50%, atleast 60%, at least 70%, at least 80%, or at least 90% endoxifen in 4hours after administration.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711 or 701, thecomposition releases in small intestine:

-   -   a. at least 10% of endoxifen after 4 hours after administration;    -   b. at least 30% of endoxifen after 6 hours after administration;    -   c. at least 40% of endoxifen after 7 hours after administration;    -   d. at least 50% of endoxifen after 8 hours after administration;    -   e. at least 50% of endoxifen after 2 hours after administration;    -   f. at least 60% of endoxifen after 2 hours after administration;    -   g. at least 70% of endoxifen after 2 hours after administration;    -   h. at least 80% of endoxifen after 2 hours after administration;        or    -   i. at least 90% of endoxifen after 2 hours after administration.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or salts thereof prepared according to any ofthe methods disclosed herein, wherein the composition is formulated fororal delivery, and wherein as tested by a method of USP 711, thecomposition is formulated to release at least 50% of endoxifen after 8hours after administration in the colon.

In another aspect, the present disclosure relates to a compositionformulated for oral administration comprising: 1 mg to 200 mg ofendoxifen per unit dose; and wherein the composition is stable for atleast 6 months.

In another aspect, the present disclosure relates to a compositionformulated for oral administration comprising: 1 mg to 200 mg ofendoxifen per unit dose;

wherein the endoxifen is at least 90% (Z)-endoxifen free base; and

wherein:

-   -   the composition has an aerobic bacterial plate count of not more        than 20,000 g/ml;    -   water content of the (Z)-endoxifen is not more than 1% as        determined by Method Ic of the USP 921;    -   residue on ignition of the (Z)-endoxifen is not more than 0.10%        as tested by a method of USP 281; or    -   a heavy metal is not more than 20 ppm as tested by Method II of        USP 231.

In another aspect, the present disclosure relates to a compositionformulated for oral administration comprising: 1 mg to 200 mg ofendoxifen per unit dose,

wherein the composition is formulated as an enteric tablet, an entericcaplet, and an enteric capsule;

wherein as determined by a method of USP 711, the composition isformulated to release in small intestine:

-   -   at least 25% of endoxifen after 4 hours after administration;    -   at least 30% of endoxifen after 6 hours after administration;    -   at least 40% of endoxifen after 7 hours after administration; or    -   at least 50% of endoxifen after 8 hours after administration; or

wherein as determined by a method of USP 711, the composition isformulated to release at least 50% of endoxifen in the colon.

In another aspect, the present disclosure relates to an oral soliddosage form composition comprising: 1 mg to 200 mg of endoxifen per unitdose;

wherein the endoxifen is at least 90% (Z)-endoxifen free base; and

wherein

-   -   the composition has an aerobic bacterial plate count of not more        than 20,000 g/ml;    -   water content of the (Z)-endoxifen is not more than 1% as        determined by Method Ic of the USP 921;    -   residue on ignition of the (Z)-endoxifen is not more than 0.10%        as tested by a method of USP 281; or    -   a heavy metal is not more than 20 ppm as tested by Method II of        USP 231; and

wherein the composition is formulated as an enteric tablet, an entericcaplet and an enteric capsule; and

wherein the endoxifen is neat; or

wherein the composition further comprises an excipient selected from thegroup consisting of a binder, a filler, a disintegrating agent, alubricant, a glidant, a control release agent, an enteric coating agent,a film forming agent, a plasticizer, a colorant, a sweetening agent, anda flavoring agent, or a combination thereof.

In another aspect, the present disclosure relates to an enteric capsule,comprising (Z)-endoxifen free base prepared by a process describedherein.

In another aspect, the present disclosure relates to an enteric capsulecomprising 1 mg to 200 mg of neat endoxifen per unit dose:

wherein the endoxifen is at least 90% (Z)-endoxifen free base; and

wherein

-   -   the composition has an aerobic bacterial plate count of not more        than 20,000 g/ml;    -   water content of the (Z)-endoxifen is not more than 1% as        determined by Method Ic of the USP 921;    -   residue on ignition of the (Z)-endoxifen is not more than 0.10%        as tested by a method of USP 281; and    -   a heavy metal is not more than 20 ppm as tested by Method II of        USP 231.

In another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the endoxifen salt isselected from the group consisting of arecoline, besylate, bicarbonate,bitartarate, butylbromide, citrate, camysylate, gluconate, glutamate,glycollylarsanilate, hexylresorcinate, hydrabamine, hydrobromide,hydrochloride, hydroxynapthanoate, isethionate, malate, mandelate,mesylate, methylbromide, methylbromide, methylnitrate, methyl sulfate,mucate, napsylate, nitrate, pamaoate (Embonate), pantothenate,phosphate/diphosphate, polygalacuronate, salicylate, stearate, sulfate,tannate, Teoclate, triethiodide, benzathine, clemizole, chloroprocaine,choline, diethylamine, diethanolamine, ethylenediamine, meglumine,piperazine, procaine, aluminum, barium, bismuth, lithium, magnesium,potassium, and zinc.

In yet another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the endoxifen salt isendoxifen gluconate.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the endoxifen salt isendoxifen gluconate selected from the group consisting of (Z)-endoxifenD-gluconate, (E)-endoxifen D-gluconate, (Z)-endoxifen L-gluconate, and(E)-endoxifen L-gluconate, or a combination thereof.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the endoxifen salt isendoxifen gluconate, and wherein the endoxifen gluconate comprises 1%,5%, 10%, 20%, 25%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%,98%, 99%, 99.5%, 99.99% or 100% (wt/wt) of (Z)-endoxifen D-gluconate or(Z)-endoxifen L-gluconate.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the endoxifen salt isendoxifen gluconate, and wherein the endoxifen gluconate comprises(Z)-endoxifen D-gluconate and (E)-endoxifen D-gluconate (wt/wt) atZ/E-ratio of 10:90, 20:80, 30:70; 40:60; 50:50, 60:40, 70:30, 80:20,90:10; 91:9; 92:8; 93:7; 94:8; 95:5, 96:4, 97:3, 98:2, 99:1; 99.5:0.5;or 99.99:0.01.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both, wherein the hormone-dependent breast disorderor the hormone-dependent reproductive tract disorder is a benign breastdisorder, hyperplasia, atypia, atypical ductal hyperplasia, atypicallobular hyperplasia, increased breast density, gynecomastia, DCIS, LCIS,breast cancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,or vulvar cancer.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both, wherein the hormone-dependent breast disorderis breast cancer.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both, wherein the hormone-dependent breast disorderis breast cancer and wherein the breast cancer is a pre-cancer, an earlystage cancer, a non-metastatic cancer, a pre-metastatic cancer, alocally advanced cancer, or a metastatic cancer.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein for the treatment and prevention of ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both, wherein the hormone-dependent breast disorderor the hormone-dependent reproductive tract disorder istamoxifen-refractory or tamoxifen resistant.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof and wherein the subject has oris at risk of having a hormone-dependent breast disorder, ahormone-dependent reproductive tract disorder, or both.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof, wherein the subject'shormone-dependent breast disorder or the hormone-dependent reproductivetract disorder is a benign breast disorder, hyperplasia, atypia,atypical ductal hyperplasia, atypical lobular hyperplasia, increasedbreast density, gynecomastia, DCIS, LCIS, breast cancer, precociouspuberty, McCune-Albright Syndrome, endometrial cancer, ovarian cancer,uterine cancer, cervical cancer, vaginal cancer, or vulvar cancer.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof, wherein the subject hasbreast cancer and wherein breast cancer is a pre-cancer, an early stagecancer, a non-metastatic cancer, a pre-metastatic cancer, a locallyadvanced cancer, or a metastatic cancer.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof, wherein the subject hasprostate cancer and wherein the subject has or is about to initiateprostate cancer therapy.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject in need thereof, wherein the subject hasprostate cancer, and wherein the subject has or is at risk of havinggynecomastia.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject that has or is at risk of having ahormone-dependent breast disorder, a hormone-dependent reproductivetract disorder, or both, and wherein the subject's hormone-dependentbreast disorder or the hormone-dependent reproductive tract disorder istamoxifen-refractory or tamoxifen resistant.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to the subject at a unit dose of 0.01 mg, 0.05 mg, 0.1 mg,0.25 mg, 0.5 mg, 0.75 mg, 1.0 mg, 1.5 mg, 2.0 mg, 4 mg, 5 mg, 10 mg, 20mg, 25 mg, 50 mg or 200 mg of (Z)-endoxifen.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to the subject at a dose of 0.01 mg, 0.05 mg, 0.1 mg, 0.25mg, 0.5 mg, 0.75 mg, 1.0 mg, 1.5 mg, 2.0 mg, 4 mg, 5 mg, 10 mg, 20 mg,25 mg, 50 mg or 200 mg of (Z)-endoxifen.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to the subject once a day, twice a day, thrice a day, fourtimes a day, every other day, twice a week, weekly, fortnightly, twice amonth, monthly, quarterly, once every six months, or annually.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein administration of thecomposition orally maintains the subject's plasma endoxifen at a levelgreater than 30 nM.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject as combination therapy.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered as a primary therapy, a neo-adjuvant therapy, or anadjuvant therapy.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject either alone in combination with a secondtherapeutic agent.

In still another aspect, the present disclosure relates to a compositioncomprising (Z)-endoxifen or a salt or polymorph thereof prepared by anyof the processes described herein, wherein the composition isadministered to a subject either alone in combination with a secondtherapeutic agent, and wherein the second therapeutic agent isbicalutamide, enzalutamide, or an anticancer drug such as trastuzumab,antineoplastic such as capecitabine (Xeloda), carboplatin (Paraplatin),cisplatin (Platinol), cyclophosphamide (Neosar), docetaxel (Docefrez,Taxotere), doxorubicin (Adriamycin), PEGylated liposomal doxorubicin(Doxil), epirubicin (Ellence), fluorouracil (5-FU, Adrucil), gemcitabine(Gemzar), methotrexate (multiple brand names), paclitaxel (Taxol),protein-bound paclitaxel (Abraxane), vinorelbine (Navelbine), eribulin(Halaven), ixabepilone (Ixempra), or inhibitors of ATP-binding cassettetransporters.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition prepared according to any one of the processes disclosedherein.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has or is at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has or is at risk of having a hormone-dependent breastdisorder, a hormone-dependent reproductive tract disorder, or both,wherein the hormone-dependent breast disorder or the hormone-dependentreproductive tract disorder is a benign breast disorder, hyperplasia,atypia, atypical ductal hyperplasia, atypical lobular hyperplasia,increased breast density, gynecomastia, DCIS, LCIS, breast cancer,precocious puberty, McCune-Albright Syndrome, endometrial cancer,ovarian cancer, uterine cancer, cervical cancer, vaginal cancer, orvulvar cancer.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has breast cancer.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has breast cancer, and wherein the breast cancer is apre-cancer, an early stage cancer, a non-metastatic cancer, apre-metastatic cancer, a locally advanced cancer, or a metastaticcancer.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has prostate cancer and wherein the subject further has oris at risk of having gynecomastia.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe subject has tamoxifen-refractory or tamoxifen resistanthormone-dependent breast disorder or hormone-dependent reproductivetract disorder.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe composition is administered to the subject at a unit dose of 0.01mg, 0.05 mg, 0.1 mg, 0.25 mg, 0.5 mg, 0.75 mg, 1.0 mg, 1.5 mg, 2.0 mg, 4mg, 5 mg, 6 mg, 8 mg, 10 mg, 20 mg, 25 mg, 50 mg, 100 mg or 200 mg.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe composition is administered once a day, twice, a day, thrice a day,four times a day, every other day, twice a week, weekly, fortnightly,twice a month, monthly, quarterly, once every six months, or annually.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinadministration of the composition orally maintains the subject's plasmaendoxifen at a steady state level greater than 30 nM.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinadministration of the composition orally achieves the subject's plasmaendoxifen at a steady state level greater than 30 nM by day 14 after theadministration of the first dose (on day 1).

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereof,wherein administration of the composition orally achieves the subject'splasma endoxifen at a steady state level greater than 30 nM by day 14after the administration of the first dose (on day 1).

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinadministration of the composition orally achieves a time to maximalplasma levels of endoxifen in the subject from 2 to 10 hours afteradministration (post-dose).

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereof,wherein administration of the composition orally achieves a time tomaximal plasma levels of endoxifen in the subject from 2 to 10 hoursafter administration (post-dose).

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereof,wherein the mean terminal elimination half-life of endoxifen in thesubject ranges from 30 to 60 hours post dose.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereof,wherein the mean terminal elimination half-life of endoxifen in thesubject ranges from 40 to 55 hours post dose.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder, a hormone-dependent reproductive tract disorder, orboth, the method comprising administering an oral composition comprising(Z)-endoxifen or a salt or polymorph thereof, wherein administration ofthe composition achieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering an oralcomposition comprising (Z)-endoxifen or a salt or polymorph thereof,wherein administration of the composition achieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancerand vulvar cancer, the method comprising administering an oralcomposition comprising (Z)-endoxifen or a salt or polymorph thereofformulated as an enteric tablet, an enteric caplet, an enteric capsule,a delayed-release tablet, a delayed-release caplet, or a delayed-releasecapsule, wherein administration of the composition achieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering an oralcomposition comprising (Z)-endoxifen or a salt or polymorph thereofformulated as an enteric tablet, an enteric caplet, an enteric capsule,a delayed-release tablet, a delayed-release caplet, or a delayed-releasecapsule, wherein administration of the composition achieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM;        and

wherein at least 50%, at least 60%, at least 70%, at least 80% or atleast 90% of the endoxifen is released in the intestines.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering an oralcomposition comprising 0.01 mg to 200 mg of (Z)-endoxifen or a salt orpolymorph thereof formulated as an enteric tablet, an enteric caplet, anenteric capsule, a delayed-release tablet, a delayed-release caplet, ora delayed-release capsule, wherein administration of the compositionachieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM;        and

wherein at least 50%, at least 60%, at least 70%, at least 80% or atleast 90% of the endoxifen is released in the intestines.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering daily an oralcomposition comprising 0.01 mg to 200 mg of (Z)-endoxifen or a salt orpolymorph thereof formulated as an enteric tablet, an enteric caplet, anenteric capsule, a delayed-release tablet, a delayed-release caplet, ora delayed-release capsule, wherein administration of the compositionachieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering daily an oralcomposition comprising 0.01 mg to 200 mg of (Z)-endoxifen or a salt orpolymorph thereof formulated as an enteric tablet, an enteric caplet, anenteric capsule, a delayed-release tablet, a delayed-release caplet, ora delayed-release capsule, wherein administration of the compositionachieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM;        and

wherein at least 50%, at least 60%, at least 70%, at least 80% or atleast 90% of the endoxifen is released in the intestines.

In another aspect, the present disclosure relates to a method oftreating a subject having or at risk of having a hormone-dependentbreast disorder or a hormone-dependent reproductive tract disorderselected from the group consisting of benign breast disorders,hyperplasia, atypia, atypical ductal hyperplasia, atypical lobularhyperplasia, increased breast density, gynecomastia, DCIS, LCIS, breastcancer, precocious puberty, McCune-Albright Syndrome, endometrialcancer, ovarian cancer, uterine cancer, cervical cancer, vaginal cancer,and vulvar cancer, the method comprising administering daily an oralcomposition comprising 0.01 mg to 200 mg of (Z)-endoxifen or a salt orpolymorph thereof formulated as an enteric tablet, an enteric caplet, anenteric capsule, a delayed-release tablet, a delayed-release caplet, ora delayed-release capsule, wherein administration of the compositionachieves:

-   -   a. a mean half-life of endoxifen in the subject ranging from 30        hours to 60 hours after administration;    -   b. a time to maximum plasma levels of endoxifen ranging from 2        hours to 10 hours after administration; and    -   c. a steady state plasma level of endoxifen greater than 30 nM;

wherein the mean area under the curve extrapolated to time infinity(AUC_(0-inf)) is 200 hr*ng/mL to 10000 hr*ng/mL, 300 hr*ng/mL to 8000hr*ng/mL, 400 hr*ng/mL to 6000 hr*ng/mL or 700 hr*ng/mL to 6000hr*ng/mL; and

wherein at least 50%, at least 60%, at least 70%, at least 80% or atleast 90% of the endoxifen is released in the intestines.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereinthe method is a primary therapy, a neo-adjuvant therapy, or an adjuvanttherapy.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereina composition is administered to a subject as combination therapy.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereina composition is administered to a subject either alone in combinationwith a second therapeutic agent.

In still another aspect, the present disclosure relates to a method oftreating a subject in need thereof comprising administering acomposition comprising (Z)-endoxifen or a salt or polymorph thereofprepared according to any one of the processes disclosed herein, whereina composition is administered to a subject with a second therapeuticagent selected from the group consisting of bicalutamide, enzalutamide,or an anticancer drug such as trastuzumab, antineoplastics such ascapecitabine (Xeloda), carboplatin (Paraplatin), cisplatin (Platinol),cyclophosphamide (Neosar), docetaxel (Docefrez, Taxotere), doxorubicin(Adriamycin), PEGylated liposomal doxorubicin (Doxil), epirubicin(Ellence), fluorouracil (5-FU, Adrucil), gemcitabine (Gemzar),methotrexate (multiple brand names), paclitaxel (Taxol), protein-boundpaclitaxel (Abraxane), vinorelbine (Navelbine), eribulin (Halaven),ixabepilone (Ixempra), and ATP-cassette binding protein transportinhibitors.

In still another aspect, the present disclosure relates to a use for thetreatment and prevention of a hormone-dependent breast disorder, ahormone-dependent reproductive tract disorder, or both, in a subject,the use comprising:

dosing the subject with a first composition comprising tamoxifen;

determining the subject's tamoxifen-metabolites profile in a test sampleobtained from the subject;

determining a reduced level of subject's plasma endoxifen based onsubject's tamoxifen-metabolites profile as to compared to a level ofplasma endoxifen in a reference tamoxifen-metabolites profile; and

administering to the subject a composition of the present disclosure.

In still another aspect, the present disclosure relates to a use for thetreatment and prevention of a hormone-dependent breast disorder, ahormone-dependent reproductive tract disorder, or both, in a subject,the use comprising:

dosing the subject with a first composition comprising tamoxifen;

determining the subject's tamoxifen-metabolites profile in a test sampleobtained from the subject;

determining a reduced level of subject's plasma endoxifen based onsubject's tamoxifen-metabolites profile as to compared to a level ofplasma endoxifen in a reference tamoxifen-metabolites profile; and

administering to the subject a composition of the present disclosure;

wherein the composition is administered orally in a dose sufficient tomaintain the subject's plasma endoxifen at a steady state level greaterthan 30 nM;

wherein the tamoxifen-metabolites profile comprises a panel of at leasttamoxifen, 4-hydroxytamoxifen, n-desmethyltamoxifen, or endoxifen;

wherein the hormone-dependent breast disorder or the hormone-dependentreproductive tract disorder is a benign breast disorder, hyperplasia,atypia, atypical ductal hyperplasia, atypical lobular hyperplasia,increased breast density, gynecomastia, DCIS, LCIS, breast cancer,precocious puberty, McCune-Albright Syndrome, endometrial cancer,ovarian cancer, uterine cancer, cervical cancer, vaginal cancer, orvulvar cancer;

wherein the hormone-dependent breast disorder is a pre-cancer, an earlystage cancer, a non-metastatic cancer, a pre-metastatic cancer, alocally advanced cancer, or metastatic cancer;

wherein the hormone-dependent breast disorder is a non-metastaticcancer;

wherein the subject has prostate cancer, and wherein the subject furtherhas or is at risk of having gynecomastia; or

wherein the hormone-dependent breast disorder or hormone-dependentreproductive tract disorder is tamoxifen-refractory or tamoxifenresistant.

In still another aspect, the present disclosure relates to a kit fortreating a subject having or at risk of having a hormone-dependentbreast disorder or hormone dependent reproductive tract disorder, in asubject in need thereof comprising: (a) a composition of the presentdisclosure; and (b) a sealed container for housing the composition; andc) instructions for use of the orally administered composition.

In still another aspect, the present disclosure relates to a kit fortreating a subject having or at risk of having a hormone-dependentbreast disorder or hormone dependent reproductive tract disorder, in asubject in need thereof comprising: (a) a composition of the presentdisclosure; and (b) a sealed container for housing the composition; andc) instructions for use of the orally administered composition, whereinthe kit comprises a second therapeutic agent selected from the groupconsisting of bicalutamide, enzalutamide and anticancer drugs such astrastuzumab, antineoplastics such as capecitabine (Xeloda), carboplatin(Paraplatin), cisplatin (Platinol), cyclophosphamide (Neosar), docetaxel(Docefrez, Taxotere), doxorubicin (Adriamycin), PEGylated liposomaldoxorubicin (Doxil), epirubicin (Ellence), fluorouracil (5-FU, Adrucil),gemcitabine (Gemzar), methotrexate (multiple brand names), paclitaxel(Taxol), protein-bound paclitaxel (Abraxane), vinorelbine (Navelbine),eribulin (Halaven), ixabepilone (Ixempra), and ATP-cassette bindingprotein transport inhibitors.

In still another aspect, the present disclosure relates to a method ofadministering a composition prepared in accordance with any of theprocesses described herein to a subject in need thereof in accordancewith instructions for use comprised in a kit comprising the composition.

EXAMPLES

Abbreviations Used Herein

ACN/PPW=Acetonitrile/Process Purified Water

NaHCO₃=Sodium hydrocarbonate

HCl=hydrochloric acid

THF=Tetrahydrofuran

MeTHF=2-Methyltetrahydrofuran

CH₂Cl₂=Dicholoromethane

EtOH=Ethanol

MeOH=Methanol

EtOAc=Ethyl acetate

IPA=Isopropyl alcohol; isopropanol

TFA=Trifluoroacetic acid

TCA=Trichloroacetic acid

PPW=Process Purified Water

IPA/PPW=Isopropyl alcohol/Process Purified Water

Example 1: Enrichment and Preparation of (Z)-Endoxifen Free Base

A. Fractional Crystallization

(Z)-Endoxifen free base (drug substance) can be prepared from a mixtureof (E)-endoxifen and (Z)-endoxifen by fractional crystallizationdescribed herein. Briefly, commercially available crude endoxifenmixture of (E)-endoxifen and (Z)-endoxifen isoforms (Astatech Ltd.China) were used as starting material to prepare (Z)-endoxifen as freebase.

i. An E/Z-endoxifen solution was prepared by suspending 1 g of endoxifenhaving (E)-endoxifen and (Z)-endoxifen at ratios (E/Z ratio) of 51/1,1/1.8, and 1/5.8, each in a 10× volume of the solvent, isopropanol(IPA). A second set of solutions was prepared by suspending 1 g each ofendoxifen at E/Z ratios of 51/1, 1/1.8, and 1/5.8 in 10× volume of thesolvent EtOAc. The solutions were heated to a first temperature of 50°C. creating a hot slurry and stirred overnight at 50° C. The hot slurrysamples were filtered using a cellulose filter pore size 0.5 μm. Themixtures were then cooled to a second temperature, room temperature 23°C. while stirring for 12 hours to effect crystallization. The aliquotsof cooled samples were filtered using a cellulose filter pore size 0.5μm. Aliquots of the solid and filtrates at each of temperatures 50° C.and 23° C. were taken and monitored by HPLC-UV to determine theirE/Z-endoxifen ratio. The (Z)-endoxifen enriched filtrates and theE-endoxifen enriched filtrates were added back to their respectivemother liquors.

HPLC-UV analyses of the samples was carried out using a Lunaphenyl-hexyl column (4.6 mm×150 mm×3 μm of particle size). The columntemperature was 40° C. and sample compartment temperature was maintainedat ambient temperature. Samples were prepared by dissolving solids andfiltrates at 0.2 mg/ml of sample in methanol as sample buffer. Sampleswere then injected into the column in an injection volume of 20 μl.Likewise, mildly acidic (pH 4.3) ammonium formate buffer/methanol in themobile phase with gradient elution was used. Mobile phase systemconsisting of: Mobile Phase A (MPA) buffer was 10 mM HCOONH₄ with 0.03%HCOOH buffer in water, and Mobile Phase B (MBP) buffer was 10 mM HCOONH₄in methanol with a gradient program used.

TABLE 1 Gradient Program Time (m) MPA (%) MPB (%) 0 50 50 15 30 70 25 595 30 50 50

The run time was 30 m, the flow rate was 1.0 mL/m, and the delay timewas 5 m. Samples were detected by UV detection at 243 nm. Retentiontimes were typically from 12.95 to 13.20 m for (E)-endoxifen and from14.0 to 14.33 m for (Z)-endoxifen. Relative Retention Time (RRT) for(E)-endoxifen was 0.4 to 0.9 and 1.0 for (Z)-endoxifen.

TABLE 2 Enrichment of Endoxifen (Z)- and (E)- isomers from hot slurry ofE/Z- endoxifen mixture in IPA and EtOAc. Sample 1 2 3 4 5 6E/Z-endoxifen 51/1 1/1.8 1/5.8 51/1 1/1.8 1/5.8 ratio E/Z-endoxifen 1 g1 g 1 g 1 g 1 g 1 g Solvent IPA (10 X) EtOAc (10 X) 50° C. Solid 85/1N/A 1/4.2 ≥100/1 1.9/1   1/24 (97.0%/1.1%) (dissolved) (18.9%/79.6%)(97.9%/0.7%) (65.4%/34.2%)  (3.9%/94.8%) Filtrate 12/1 N/A 1/47     5.4/1 1/2.4 1/2.5 (87.0%/7.0%) (dissolved)  (2.1%/97.4%)(66.2%/12.3%) (29.2%/68.7%) (28.0%/70.1%) 23° C. Solid ≥100/1     1/2.11/90  ≥100/1 1.3/1   1/6.9 (98.2%/0.3%) (31.8%/67.2%)  (1.1%/98.1%)(99.0%/0.4%) (55.9%/43.1%) (12.6%/86.9%) Filtrate 3.6/1  1/1.7 1/1.3    1.6/1 1/6.7 1/2.6  (70.1%/19.6%) (36.5%/60.6%) (43.4%/54.9%)(37.7%/23.0%) (12.6%/84.3%) (26.2%/84.3%)

Results (Table 2) show that Z-endoxifen can be enriched using acommercial crude E/Z-endoxifen starting material from a hot slurry ofthe endoxifen mixtures made with IPA and EtOAc. Z-endoxifen can bepurified from E/Z-endoxifen mixtures having E/Z at 1:1 and with higherZ-endoxifen levels in the mixture, for example, 1:8, and 1:5.8.Substantial Z-endoxifen enrichment was achieved in a single step usingfractional crystallization method, for example with an E/Z ratio of 1:90described herein Example 1A. (Z)-endoxifen was seen to be more solublethan (E)-endoxifen in EtOAc. (Z)-endoxifen also tended to remain in thefiltrate in greater amounts in EtOAc as compared with IPA. Both solidsand filtrates are useful for the preparation of (Z)-endoxifen free base.Serial enrichment of Z-endoxifen by adding filtrates back to the firstfiltrate (first mother liquor) can also be performed.

Similarly, E-endoxifen can be purified from E/Z-endoxifen mixture havinghigher E-endoxifen levels in the E/Z mixture.

ii. Similar results were obtained in experiments where an E/Z-endoxifenmixture was suspended in a hot slurry prepared with a mixture of IPA/PPW(1:1 v/v) and extracted and washed with precooled IPA/PPW (1:1) to formcrystalline solids and filtrates with enriched (Z)-endoxifen.

B. Recrystallization from Solid Fraction

Serial enrichment of (Z)-endoxifen can be performed by subjecting theendoxifen mixture with E/Z-endoxifen ratio of 51:1, 1:1.8 and 1:5.6, andthe solids and mother liquors obtained from the fractionalcrystallization to one or more rounds of recrystallization.

The (Z)-endoxifen can be enriched in substantially purified form fromcrude E/Z-endoxifen mixtures, (Z)-enriched crystalline solids and(Z)-enriched mother liquors by recrystallization with solvents such asethanol, ethyl acetate/n-heptane, Acetone/MTBE, IPA, IPA/PPW, and thelike under the conditions as described herein.

Ethanol.

Crystalline solids obtained from E/Z-endoxifen hot IPA slurry mixtureabove were used to obtain further purified Z-endoxifen. After cooling to23° C., the solid fractions of sample 2 (80 g) and sample 3 (79 g) ofTable 2 above were combined and recrystallized using 1000 mL (1 L) EtOH.The ethanol mixture was heated to reflux. The dissolution point was at75° C. The heated solution was filtered using a cellulose filter (poresize 0.5 μm) to remove foreign impurities such as cotton fibers bypolish filtration. The filtrate solution was then stirred initially at70° C. for 30 minutes. The solution was then cooled to 0° C.-5° C. andstirred continually for 5 h at 0° C.-5° C. The solution was filteredagain. The crystals on the filter were washed with 200 mL of EtOH.

Results show that solid crystals of endoxifen had E/Z ratio of ≥1/100(0.22%/99.48%) and endoxifen in the filtrate had E/Z ratio of 1/5.9(13.93%/82.48%) when recrystallized with EtOH. HPLC analysis of at least3 different experiments conducted by this method showed that (Z)endoxifen retention time was 14.00 m, E-endoxifen retention time was13.14 m. (Z)-endoxifen levels ranged from 97% to 99.48% while(E)-endoxifen levels ranged from 0.22% to 2.14%. Other impurities were<1%.

Ethyl Acetate/n-heptane.

Feasibility of recrystallization with EtOAc/n-heptane was studied asdescribed below.

5 g Endoxifen (E/Z ratio 1/5.8) was added to EtOAc/n-heptane (140 mL,1/1 v/v) and heated to reflux (˜82° C.) while stirring overnight toobtain a clear solution. The cloud point of the mixture was 30° C. Themixture was then cooled to room temperature to precipitate the productand filtered. After isolation as described above, endoxifen of the solidproduct was determined by HPLC to have an E/Z ratio of 1/8.1(10.9%:88.60%) with a product yield of 66% (3.3 g). (Z)-Endoxifen in thefiltrate was enriched with E/Z ratio of 1:3 (23.94%:74.47%). Therecrystallization using EtOAc/n-heptane can also provide purifiedZ-endoxifen in ratios similar to the hot IPA slurry as shown in Table 3.

TABLE 3 Recrystallization of (Z)-endoxifen from E/Z-endoxifen mixtureusing EtOAc/n-heptane solvent. Parameter Result E/Z endoxifen—E/Z ratio1/5.8  5 g EtOAc/n-heptane solvent (1:1) 140 mL Heat to dissolution ~82°C. (reflux) Cloud point ~30° C. Cool to RT and filtration   23° C., 1 hrE/Z ratio of the product Solid 3.3 g, 1/8.1 (10.90%/88.60%) Filtrate1/3.0 (23.94%/72.47%)

Acetone/MTBE.

0.5 g Endoxifen (E/Z ratio 1/1.8) was added to Acetone (10 v; 5 mL)solvent, and antisolvent MTBE (9 mL) was added to the solution andheated to reflux (˜82° C.) while stirring overnight to obtain a clearsolution. The cloud point of the mixture was ˜22° C. The mixture wasthen cooled to room temperature to precipitate the product and filtered.Endoxifen in the solid product was determined by HPLC to have an E/Zratio of 39/1 while the filtrate was enriched with (Z)-endoxifen withE/Z ratio of 1:2.6. Acetone/MTBE solvent system is useful to separateE-endoxifen to solid from the (Z)-endoxifen in the filtrate.

C. Recrystallization from Mother Liquor

Purified (Z)-endoxifen can be recovered from mother liquor enriched with(Z)-endoxifen as shown in the following Tables 4 and 5.

i. Starting material of crude E/Z-endoxifen mixture can be dissolved ina first solvent, ethyl acetate, to form a first mother liquor from which(Z)-endoxifen can be recovered by a second step of addition of a secondsolvent such as n-heptane or ethanol as shown in Table 4.

TABLE 4 Recrystallization of Z-endoxifen Results Lot Number Parameter 12 E/Z-endoxifen (E/Z = 1/1.8)  5 g 10 g EtOAc (First solvent) 50 mL 50mL Heat to dissolution ~68° C. ~78° C. (reflux) Cloud Point° ~33° C.~38° C. Cool to RT and filtration   23° C., 1 h   23° C., 1 h E/Z ratioof product Solid 0.87 g, 8.5/1 1.53 g, 1/1 (89.37%/10.49%)(49.32%/49.80%) Filtrate 1/3.8 1/2.6 (20.13%/76.86%) (27.29%/70.09%)Filtrate concentration To 5 mL, 25 mL To dryness Heat to dissolution~68° C. ~68 C. Add solvent/anti-solvent n-heptane, 40 mL, EtOH, 50 mL,(second solvent) no precipitation heat to dissolution observed Cool toRT and filtration   23° C.,   23° C. overnight (cloud point ~28 C.) E/Zratio of product Solid 3.0 g, 1/4.2 3.2 g, 1/12 (18.70%/78.95%)(7.52%/90.24%) Filtrate 1/3.2 1/1.5 (21.65%/70.35%) (39.35%/57.76%)

ii. (Z)-endoxifen was prepared by a process involving: a first step ofdissolving crude E/Z-endoxifen mixture in a first solvent, ethylacetate, enriching (Z)-endoxifen to first mother liquor; a second stepof addition of a second solvent, IPA to form crystalline (Z)-endoxifenand a second mother liquor; and a step of addition of a third solventethanol to recrystallize and recover (Z)-endoxifen from the secondmother liquor.

Briefly, endoxifen (E/Z=1/1.8, 30 g) and (E/Z=1.2/1, 50 g non-GMP) wasdissolved in EtOAc 300 mL and 500 mL respectively, heated to dissolution(˜70° C.). The mixture was then cooled to 25° C. while stirringovernight for precipitation. The suspension was filtered and E/Z ratioof solid and the filtrate was determined by HPLC.

Next, the filtrates were concentrated to dryness and IPA 230 mL and 900mL respectively were added to form hot slurries (50° C.) while stirring.The hot slurries were continuously stirred at 50° C. overnight. Theslurries were then cooled to 25° C. for 0.5 h and 1 h respectively, andfiltered. Z-endoxifen in the solid and the filtrates were determinedagain.

The solids of each sample (14.8 g and 21.6 g) were furtherrecrystallized from 148 mL and 220 mL EtOH by heating to dissolution atNMT 70° C. and cooling to 25° C. for 0.5 h and 1 h respectively. Thesamples were further cooled to 0° C.-5° C. for 1 h and filtered. The E/Zratio of the solid and the filtrates were again determined by HPLC. E/Zratios of each sample and purity are provided below in Table 5.

TABLE 5 Recovery of (Z)-endoxifen from (Z)-endoxifen enriched motherliquor Results Lot Number Parameter 1 2 E/Z-endoxifen ratio 1/1.8 1.2/1E/Z-endoxifen amount 30 g 50 g EtOAc 300 mL 500 mL Heat to Dissolution~70° C. ~70° C. Cloud Point ~33° C. ~33° C. Cool to RT 25° C., 3 h 25°C., overnight E/Z ratio of Solid 4.5/1 (80.52%/17.73%) 13.5/1(92.44/6.83%) product Filtrate 1/5.0 (16.02%/80.71%) 1/6.8(12.34%/84.45%) Filtrate concentration To remove EtOAc Swap with IP (400ml X 2) Add IPA 230 mL 900 mL Final Volume 250 mL Hot Slurry 50° C.overnight 50° C., overnight Cool to RT and filtration 25° C., 4 h 25°C., 4 h E/Z ratio of Solid 1/31.7 (3.0%/95.15%) 1.31.9 (2.94%/93.74%)product Filtrate 1/1.6 (36.16%/58.78%) 1/1.7 (32.17%/55.92%)E/Z-endoxifen for 14.8 g 21.6 g recrystallization EtOH 148 mL 220 mLHeat to Dissolution ~70° C. ~70° C. Cool to RT 25° C., 0.5 h 25 C., 1 hE/Z ratio of Solid — 1/76.9 (1.26%/91.91%) Product Filtrate — 1/20.1(4.57%/91.91%) Cool to 0°C-5°C 0° C.-5° C., 1 h 0° C.-5° C., 1 h andfiltration E/Z ratio of Solid (?) 1/51.8 (1.86%/96.31%) 1/118.7(0.82%/97.30%) product Filtrate (?) 1/6.8 (12.72%/86.23%) 1/8(10.54%/84.66%) Total Yield 31% 29%

Results show that purified crystalline (Z)-endoxifen of greater than 90%can be obtained by subjecting an E/Z-endoxifen mixture to fractionalcrystallization with a first solvent, ethyl acetate, to form crystallinesolid and a first mother liquor, subjecting the first mother liquor tocrystallization with a second solvent such as hot IPA to form a secondcrystalline solid and second mother liquor, and then recrystallizationof the solids from a third solvent such as ethanol. A yield of(Z)-endoxifen of greater than 95% is also feasible. The ratio of(E)/(Z)-endoxifen can be improved from ˜1.2:1 and 1:1.8 to 1:51 to over1:118. The impurity profile of (Z)-endoxifen was analyzed by an HPLCmethod as described above and was found to be less than <2%. Retentiontime of (Z)-endoxifen and (E)-endoxifen were 14.19 m and 13.2 mrespectively.

Example 2: Reequilibriation of (E)-Endoxifen-Rich E/Z-Mixture

(E)-endoxifen-rich E/Z-endoxifen mixture is also useful for thepreparation Z-endoxifen free base (in addition to being useful for thepreparation of purified (E)-endoxifen). (E)-endoxifen-rich E/Z-endoxifenmixture is reequilibriated from a E/Z ratio of ≥1.5:1 (for example, E/Zratio of 51:1) to about ˜1:1 mixture. The present disclosure embracesthe idea that reequilibration may be performed with a startingE/Z-endoxifen mixture having any E/Z ratio, such as 99:1 to 1:99. Suchreequilibration is useful for preparation of stable (E)/(Z)-endoxifen˜1:1 mixture as well as further enrichment and purification ofZ-endoxifen free base as described in Example 1 above.

Briefly, (E)-endoxifen is converted to (Z)-endoxifen by dissolving crudeE/Z-endoxifen in an inert solvent such as acetonitrile/water (8:1) ordichloromethane in the presence of an acid such as hydrochloric acid,trichloroacetic acid or trifluoroacetic acid. The reaction mixture withACN/PPW was stirred at 60° C. for 1 hour while the reaction mixture withdichloromethane and TFA was stirred at RT for 1 hour. After work up wasperformed as described in Table 6, the E/Z ratio of the product wasdetermined to be E/Z-endoxifen (ratio about 1:1) with a purity greaterthan 90%.

TABLE 6 Reequilibrium of E/Z-endoxifen (E/Z ratio 51/1) to E/Z ratio ofabout 1:1 Results Sample Parameter 1 2 E/Z-endoxifen 0.5 g 0.5 g (E/Zratio = 51/1) (off-white powder (off-white powder) Solvent ACN/PPW =8/1, 5 mL CH₂Cl₂, 5 mL (white slurry) (White slurry) Acid 6N HCl, 5 mLTFA, 5 mL ~60° C., 6 h ~23° C., 1 h Stir (Pink solution to clear (orangeclear solution) solution) Work up Quench with saturated Quench withsaturated solution of NaHCO₃, solution of NaHCO₃, wash the organic washthe organic layer with brine layer with brine and organic layer is andorganic layer is evaporated to dryness evaporated to dryness ProductBeige solid Off-white solid E/Z ratio ~1/1 (49.28%/47.70%) ~1.3/1(54.51%/41.9%) Purity 96.98% 96.20%

Example 3: Purification of (E)-Endoxifen

(E)-endoxifen was similarly purified by recrystallization in EtOH (500mL) by heating the reaction mixture to NMT 70° C. to get a clearsolution, cooled to 0° C.-5° C. to precipitate the product. Afterpurification by methods described herein, 38.1 g of (E)-endoxifen wasobtained in 76% yield with 97.6% purity.

Example 4: Scale Up

Synthetic and purification methods disclosed herein also are useful inscaling up the preparation of (Z)-endoxifen for making industrial drugproduct dosage forms, for example, oral, transdermal/topical, nasal,intraductal parenteral dosage forms, for use clinical use in subjects.Feasibility studies of crystallization using 5 g, 100 g, and 110 g ofE/Z-endoxifen mixture, compounds of Formula (III) at an E/Z ratio of1/5.8 were performed by suspending the E/Z-endoxifen mixtures in 10×volume of IPA and heated to 50° C. and stirred for 4 h at 50° C. Thesamples were filtered using a cellulose filter pore size 0.5 μm and theE/Z ratio was determined in the solids. Following filtration, thesamples were cooled to 23° C. and stirred continuously for 4 h at 23° C.The samples were filtered again. Solids and the filtrates were monitoredfor (E)-endoxifen and (Z)-endoxifen levels by HPLC as described aboveand the results are provided below in Table 7.

TABLE 7 Scale up Preparation of (Z)-endoxifen Results Sample Parameter 12 3 E/Z- 5 g 100 g 110 g endoxifen ratio 1/5.8 IPA (10 X)  50 mL  1000mL  1100 mL 50° C. Solid  1/20 1/22  1/16  (4.71%/94.50%) (4.25%/94.88%)(5.85%/93.24%) Fil- 1/3.1 1/2.2 1/2.3 trate  (26.54%/72.23%) (30.62%/67.52%) (29.94%/68.04%) Cooling Solid 1/78 1/78  1/74  to(1.26%/97.69%) (2.14%/97.24%) (1.32%/98.12%) 23° C. Fil-  1/1.1 1.1/1   1.2/1    stirring trate  (50/0.75%/47.1%)  (51.47%/45.88%) (53.03%/43.81%) for 4 h

Results show that Z-endoxifen can be obtained from an E/Z-endoxifenmixture from IPA solvent at concentrations higher than ≥90% ascrystalline solid as well as ≥65% (Z)-endoxifen free base in thefiltrate. IPA is thus useful as a first solvent, or second solvent orboth.

Example 5: Industrially Scalable Preparation of[4-[2-(methylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone)

A suitable 10 L reactor was charged with starting material[4-[2-(dimethylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone, thecompound of Formula (I) (0.5 kg, 1.0 equiv.), DIPEA (1.5 kg, 6.65equiv., 3.0 wt./wt.) and tetrahydrofuran (5 L, 10 vol./wt., 8.9 wt./wt.)under N₂ atmosphere. 1-Chloroethyl chloroformate (1.7 kg, 6.65 equiv.,3.3 wt./wt.) was added slowly while maintaining the internal temperatureat NMT 20° C. The mixture was heated to reflux and stirred at reflux forNLT 12 hr. The mixture was evaporated under reduced pressure at NMT 75°C. until the volume reached the lowest agitateable volume. Methanol (2.5L, 5 vol./wt., 4.0 wt./wt.) was added slowly and the mixture wasdistilled under reduced pressure at NMT 75° C. until the volume reachedthe lowest agitateable volume. Methanol (2.5 L, 5 vol./wt., 4.0 wt./wt.)was added and the mixture was distilled under reduced pressure at NMT75° C. until the volume reached the lowest agitateable volume. Methanol(2.5 L, 5 vol./wt., 4.0 wt./wt.) was once again added and the mixturewas further distilled under reduced pressure at NMT 75° C. until thevolume once again reached the lowest agitateable volume. Methanol (2 L,4 vol./wt., 3.2 wt./wt.) and 6N HCl (2 L, 4 vol./wt., 4.0 wt./wt.) wereadded to the mixture and the mixture was heated to reflux. The mixturewas stirred at reflux for NLT 12 hr. After reaction completion, themixture was evaporated under reduced pressure at NMT 75° C. until mostof MeOH was removed. The mixture was cooled to 25±5° C. and 8N NaOH (2.5L, 5 vol./wt., 5.0 wt./wt.) was added slowly until the pH of the mixturewas 11-12. The mixture was stirred at 0-5° C. for NLT 2 hr. The mixturewas filtered and washed with H₂O (1 L, 2 vol./wt.) and ethyl acetate (1L, 2 vol./wt., 1.8 wt./wt.). The wet cake was dried at NMT 50° C. underreduced pressure to afford(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, thecompound of Formula (II). Yield: 297 gm, 63%; Purity: 100% (ExpectedYield: 60-90%)

Example 6: Industrially Scalable Preparation of[4-[2-(methylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone)

A suitable 2 L reactor was charged with starting material[4-[2-(dimethylamino)ethoxy]phenyl](4-hydroxyphenyl)methanone, thecompound of Formula (I) (70 gm, 1.0 equiv.), N-Ethyldiisopropylamine(126 gm, 4.0 equiv., 1.8 wt.) and tetrahydrofuran (700 mL, 10.0 vol.,8.9 wt.) under N₂ atmosphere. The mixture was heated to NLT 60° C. and1-chloroethyl chloroformate (140 gm, 4.0 equiv., 2.0 wt.) added. Themixture was heated to reflux and stirred for NLT 12 hr. The mixture wasconcentrated until the volume reached 3 vol. Methanol (350 mL, 5.0 vol.,4.0 wt.) was added slowly and the mixture was concentrated until thevolume reached 3 vol. (210 mL). Next, methanol (350 mL, 5.0 vol., 4.0wt.) was added and the mixture was concentrated until the volume reaches4 vol. (280 mL). 6N HCl (280 mL, 4.0 vol., 4.0 wt.) was added and themixture was heated to reflux. The mixture was stirred at reflux for NLT12 hr. After reaction completion, the mixture was concentrated until thevolume reaches ˜4 vol. (280 mL). The mixture was cooled to ambienttemperature, and 8N NaOH (NLT 350 mL, 5.0 vol., 5.0 wt.) was addedslowly until the pH of the mixture is NLT 13. Ethyl acetate (280 mL, 4.0vol., 3.6 wt.) was added to the mixture for extraction. After phaseseparation, to the aqueous layer was added 6N HCl (NLT 42 mL, 0.6 vol.,0.6 wt.) until the pH was 8-10. The mixture was cooled to 0±5° C. andstirred for NLT 2 hr. The mixture was filtered and washed with purifiedwater (NLT 140 mL, 2.0 vol., 2.0 wt.) and ethyl acetate (NLT 140 mL, 2vol., 1.8 wt.). The wet cake was dried at NMT 60° C. under reducedpressure to afford(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, thecompound of Formula (II). Yield: 55.2 gm, 83%; Purity: 100% (ExpectedYield: NLT 70%).

Example 7: Industrially Scalable Preparation of E/Z-Endoxifen Mixture ina McMurry Reaction

A suitable 10 L reactor was charged with Zn powder (0.27 kg, 4.0 equiv.,0.9 wt./wt.) and tetrahydrofuran (1.5 L, 5 vol./wt., 4.4 wt./wt.) underN₂ atmosphere. TiCl₄ (0.42 kg, 2.0 equiv., 1.4 wt./wt.) was added slowlywhile maintaining the internal temperature at NMT 15° C. The reactionwas heated to reflux and stirred at reflux for NLT 2 hr. A suspension of(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, thecompound of Formula (II) obtained from Step 1, (0.297 kg, 1.0 equiv.)and propiophenone (0.21 kg, 1.5 equiv., 0.7 wt./wt.) in tetrahydrofuran(3.0 L, 10 vol./wt., 8.9 wt./wt.) was added and the reflux was continuedfor NLT 8 hr. The mixture was cooled to 20-30° C. and added into 25%ammonium chloride (5.9 L, 20 vol./wt., 20 wt./wt.)/silica (diatomaceousearth/silicon dioxide; tradename Celite® S) available from Sigma Aldrich(0.3 kg, 1.0 wt./wt.) mixture. The mixture was filtered and washed withtetrahydrofuran (0.9 L, 3 vol./wt., 2.7 wt./wt.).

The mixture was settled for phase separation. The organic layer wascollected and the aqueous layer was washed with tetrahydrofuran (0.9 L,3 vol./wt., 2.7 wt./wt.). The organic layer was again collected andcombined with the first organic layer. The combined organic layer waswashed with 40% K₂CO₃ (1.2 L, 4 vol./wt., 5.6 wt./wt.). The organiclayer was concentrated at NMT 75° C. under reduced pressure until thevolume reached the lowest agitateable volume. Ethyl acetate (1.5 L, 5vol./wt., 4.5 wt./wt.) was added and the mixture was distilled underreduced pressure at NMT 75° C. until the volume reached the lowestagitateable volume. Ethyl acetate (1.5 L, 5 vol./wt., 4.5 wt./wt.) wasadded and the mixture was distilled under reduced pressure at NMT 75° C.until the volume reached 5 vol. (1.5 L). The mixture was heated todissolution and n-heptane (3.0 L, 10 vol./wt., 6.8 wt./wt.) was added.The mixture was cooled to 0±5° C. and stirred at 0±5° C. for NLT 2 hr.The mixture was filtered and the residue was washed with ethylacetate/n-heptane=1/2 (v/v, 1.5 L, 5 vol./wt., 3.7 wt./wt.). The residuewet cake was dried under reduced pressure at NMT 60° C. to afford amixture of (Z)-endoxifen and (E)-endoxifen,(E/Z)-4-[1-[4-[2-(Methylamino)ethoxy]phenyl]-2-phenyl-1-buten-1-yl]-phenol,compounds of Formula (III). Yield: 176 g, 42%; Purity: 81.96%. E/Zratio: 3.1:1 (Expected yield—40-60%).

A suitable 10 L reactor was charged with the above (E/Z)-endoxifen(0.250 kg, 1.0 wt.), isopropanol (1.25 L, 5 vol./wt., 3.9 wt./wt.) andpurified water (1.25 L, 5 vol./wt., 5.0 wt./wt.). The mixture was heatedto 70±5° C. and stirred at 70±5° C. for NLT 2 hr. The mixture was cooledto 0±5° C. and stirred at 0-5° C. for NLT 2 hr. The mixture was filteredand washed with pre-cooled isopropanol/purified water=1/1 (0.5 L, 2vol./wt., 3.6 wt./wt.). The residue wet cake was dried at NMT 60° C. toafford a mixture of (Z)-endoxifen and (E)-endoxifen,(E/Z)-4-[1-[4-[2-(Methylamino)ethoxy]phenyl]-2-phenyl-1-buten-1-yl]-phenol,compounds of Formula (III). Yield: 105 g, 41%; Purity: 96.79%. E/Zratio: 48.6:1 (Expected yield—40-60%).

Example 8: Industrially Scalable Preparation of E/Z-Endoxifen Mixture ina McMurry Reaction

To a suitable reactor was charged zinc powder (10 gm, 4.0 equiv., 1.0wt.) and tetrahydrofuran (50 mL, 5.0 vol., 4.3 wt.) under N₂ atmosphere.Titanium(IV) chloride (14 gm, 2.0 equiv., 1.4 wt.) was added slowlywhile maintaining the internal temperature at NMT 45° C. The reactionwas heated to 65±5° C. and stirred for NLT 2 hr. A suspension of(4-hydroxyphenyl)(4-(2-(methylamino)ethoxy)phenyl)methanone, thecompound of Formula (II) obtained from Example 5, (10 gm, 1.0 equiv.)and propiophenone (5 gm, 1.0 equiv., 0.5 wt.) in Tetrahydrofuran (100mL, 10.0 vol., 8.5 wt.) was added to the above mixture and stirred at65±5° C. for NLT 8 hr. The mixture is cooled to NMT 30° C. and 40% K₂CO₃(20 mL, 2.0 vol., 2.0 wt.) is added. The mixture was stirred for NLT 1hr. The mixture was filtered and washed with Me-THF (40 mL, 4.0 vol.,3.5 wt.). The filtrate was added to 40% K₂CO₃ (30 mL, 3.0 vol., 3.0 wt.)and stirred for NLT 30 min. The mixture was filtered and washed withMe-THF (20 mL, 2.0 vol., 1.7 wt.). The filtrate was settled for phaseseparation and the organic layer is concentrated until the volumereached 3 vol. (30 mL). The mixture was extracted with 1N NaOH (100 mL,10.0 vol., 10.0 wt.) and the organic layer was discarded. The aqueouslayer was added sodium chloride (1 gm, 0.10 wt.) and then extracted 4times with Me-THF (50 mL, 5.0 vol., 4.3 wt.). The 4 organic layers arecombined and concentrated until the volume reached 5 vol. The mixture isextracted with 20% NaCl (50 mL, 5.0 vol., 5.0 wt.). The organic layerwas added isopropanol (50 mL, 5.0 vol., 3.9 wt.) and the mixture wasconcentrated until the volume reached 3 vol. isopropanol (50 mL, 5.0vol., 4.5 wt.) was added and the mixture is concentrated until thevolume reached 3 vol. (30 mL). The mixture was heated to 65±5° C. andn-Heptane (120 mL, 12.0 vol., 8.2 wt.) was added. The mixture was cooledto NMT 5° C. and stirred at NMT 5° C. for NLT 2 hr. The mixture wasfiltered and washed with n-Heptane (50 mL, 5 vol., 3.4 wt.). The wetcake was dried at NMT 70° C. under reduced pressure to afford a mixtureof (Z)-endoxifen and (E)-endoxifen,(E/Z)-4-[1-[4-[2-(Methylamino)ethoxy]phenyl]-2-phenyl-1-buten-1-yl]-phenol,compounds of Formula (III). Yield: 6.7 g, 45%; Purity: 96.79%. E/Zratio: 48.6:1 (Expected yield—NLT 30%).

Example 9: Industrially Scalable Enrichment of Z-Endoxifen

A suitable reactor was charged with the mixture of (Z)-endoxifen and(E)-endoxifen, compounds of Formula III, (0.105 kg, 1.0 wt.) and firstsolvent, ethyl acetate, (1.1 L, 10 vol./wt., 9.0 wt./wt.). The mixturewas cooled to 0-5° C. and 6N HCl (0.3 L, 3 vol./wt., 3.0 wt./wt.) wasadded slowly to the mixture. The mixture was heated to 60±5° C. andstirred at 60±5° C. for NLT 6 hr. The mixture was cooled to 0±5° C. and8N NaOH (0.3 L, 3.0 vol./wt., 3.0 wt./wt.) was added slowly until the pHof the mixture was ≥12.

The mixture was settled for phase separation, the organic layer wascollected and the aqueous layer was washed with the first solvent, ethylacetate (0.5 L, 5 vol./wt., 4.5 wt./wt.). The organic layer wascollected. The combined organic layers were washed with 20% NaCl (0.3 L,3 vol./wt., 3.0 wt./wt.). The organic layer was treated with activatedcarbon (charcoal) (0.005 kg, 0.05 wt.) and stirred at 50±5° C. for NLT 1hr. The mixture was filtered through a bed of diatomaceous earth/silica(Celite® S) and washed with ethyl acetate (0.5 L, 5 vol./wt., 4.5wt./wt.). The filtrate was concentrated at NMT 75° C. until the volumereached 10 vol. (1.1 L). The mixture was cooled to 0-5° C. and stirredat 0-5° C. for NLT 2 hr. The mixture was filtered and the filtrate wascollected. The filtrate was concentrated at NMT 75° C. until the volumereached the lowest agitateable volume.

A second solvent, isopropanol, (1.1 L, 10 vol./wt., 7.9 wt./wt.) wasadded and the mixture was concentrated at NMT 75° C. until the volumereached the lowest agitateable volume. Isopropanol (1.1 L, 10 vol./wt.,7.9 wt./wt.) was again added and the mixture was concentrated at NMT 75°C. until the volume reached the lowest agitateable volume. Isopropanol(1.1 L, 10 vol./wt., 7.9 wt./wt.) was added and the mixture wasconcentrated at NMT 75° C. until the volume reached 5 vol. (0.5 L). Thesuspension was stirred at 50±5° C. for NLT 6 hr. The mixture was cooledto 20±5° C. and stirred at 20±5° C. for NLT 4 hr. The mixture wasfiltered and washed with isopropanol (0.2 L, 2 vol./wt., 1.6 wt./wt.).The wet cake was dried at NMT 60° C. to afford purified (Z-endoxifen)solid compound of Formula (IV) as an off-white solid. Yield: 39.5 g,36%, (E)/(Z) ratio: 1/12.2; Purity (Z): 89.59% by HPLC, Expected Yield(20-40%).

A suitable 1 L reactor was charged with the above Z-endoxifen (39 g)solid compound of Formula (IV) and a third solvent, isopropanol (390 mL,10 vol./wt.). The suspension was stirred at 50±5° C. for NLT 6 hr. Themixture was cooled to 20±5° C. and stirred at 20±5° C. for NLT 4 hr. Themixture was filtered and washed with isopropanol (78 mL, 2 vol./wt.).The wet cake was dried at NMT 60° C. to afford purified (Z-endoxifen)solid compound of Formula (IV) as an off-white solid. Yield: 25 g, 61%,(E)/(Z) ratio: 1/55.1; Purity (Z): 95.81%, Expected Yield (50-70%).

In this manner, polymorph Form I of endoxifen was prepared. XRPDpatterns for two batches of the resultant polymorph are provided inFIGS. 9 and 10. The XRPD peaks from FIGS. 9 and 10 are provided in Table8.

TABLE 8 XRPD peaks from FIGS. 9 and 10. FIG. 9 FIG. 10 Peak PositionRelative Peak Position Relative (°2θ) Intensity (%) (°2θ) Intensity (%)7.1 15 7.1 8 9.3 22 9.3 13 12.4 8 12.3 4 14.2 26 14.2 16 15.2 14 15.2 1016.0 20 16.0 14 16.8 58 16.8 49 17.1 100 17.1 100 18.2 40 18.2 35 18.615 18.6 8 18.9 12 18.8 8 19.8 14 19.9 6 20.9 32 20.9 31 21.3 24 21.3 1921.7 53 21.8 55 22.5 15 22.5 9 23.2 19 23.2 9 24.3 30 24.4 29 25.3 1525.3 12 26.5 13 26.5 11 26.8 20 26.9 23 28.0 9 28.0 7 29.0 9 29.0 9

Example 10: Industrially Scalable Enrichment of ˜1:1E/Endoxifen/Z-Endoxifen Mixture

A suitable reactor was charged with the mixture of (Z)-endoxifen and(E)-endoxifen, compounds of Formula III, (0.420 kg, 1.0 wt.) and ethylacetate (4.2 L, 10 vol./wt., 9.0 wt./wt.). The mixture was cooled to0-5° C. and 6N HCl (1.3 L, 3 vol./wt., 3.0 wt./wt.) was added slowly.The mixture was heated to 60±5° C. and stirred at 60±5° C. for NLT 6 hr.The mixture was cooled to 0±5° C. and 8N NaOH (1.3 L, 3.0 vol./wt., 3.0wt./wt.) was added slowly until the pH of the mixture was ≥12.

The mixture was settled for phase separation, the organic layer wascollected and the aqueous layer was washed with ethyl acetate (2.0 L, 5vol./wt., 4.5 wt./wt.). The organic layer was collected. The combinedorganic layers were washed with 20% NaCl (1.3 L, 3 vol./wt., 3.0wt./wt.). The organic layer was treated with activated carbon (charcoal)(0.02 kg, 0.05 wt.) and stirred at 50±5° C. for NLT 1 hr. The mixturewas filtered through diatomaceous earth/silica (Celite® S) bed andwashed with ethyl acetate (2.0 L, 5 vol./wt., 4.5 wt./wt.). The filtratewas concentrated at NMT 75° C. until the volume reached 5 vol. (2.0 L).The mixture was heated to reflux and then cooled to 50±5° C. n-Heptanewas added slowly at 50±5° C. The mixture was cooled to 0±5° C. andstirred at 0±5° C. for NLT 2 hr. The mixture was filtered and washedwith EtOAc/n-Heptane=1/2 (v/v). The wet cake was dried under reducedpressure at NMT 60° C. to afford substantially pure ˜1:1 E/Z-endoxifen.[Yield: 106 g, 25%; (E)/(Z) ratio: 1.1/1; Purity (E/Z): 97.94%, ExpectedYield (20-40%)].

In this manner, polymorph Forms II and III of endoxifen were prepared.Detailed parameters for the preparation of polymorph Forms II and IIIare provided in Table 9, and XRPD patterns are provided in FIGS. 11-13.The XRPD peaks from FIGS. 11-13 are provided in Table 10

TABLE 9 Parameters for isolation of polymorph Form II and Form III Entry1 2 3 Reactor capacity 10 L 10 L 50 L E/Z-endoxifen input 0.37 kg 0.17kg 0.83 kg 6N HC1 1.1 kg (3 vol.) 0.68 kg (4 vol.) 3.32 kg (4 vol.)Reaction time 60° C., 6 hr 60° C., 6 hr 60° C., 6 hr E/Z ratio in IPC1.1 1.1 1.1 pH after 8N NaOH 14 13 14 neutralization NaCl extraction 20%NaCl, 1.1 kg 20% NaCl, 1.0 kg 5% NaCl, 2.5 kg Org. layer concentrate to1.9 L (5.1 vol.) to 1 L (5.8 vol.) to 4 L (4.8 vol.) to 5 vol. n-Heptane(10 vol.) 2.5 kg, 40 min 1.2 kg, 30 min 5.6 kg, 36 min addition at 50°C. Agitation at 50° C. 150 rpm, 40 min 130 rpm, 1 hr 150 rpm, 2 hr Coolto 0° C. 2 hr 4 hr 5 hr Agitation at 0° C. 150 rpm, 2 hr 140 rpm, 6 hr150 rpm, 6 hr Filtration time 10 min 10 min 20 min Drying 50° C., 4 hr68° C., 18 hr 69° C., 26 hr E/Z ratio of API 1.1 0.92 1.1 pattern FormII (FIG. 11) Form III (FIG. 13) Form II (FIG. 12)

TABLE 10 XRPD peaks from FIGS. 11, 12 and 13. FIG. 11 FIG. 12 FIG. 13Peak Relative Peak Relative Peak Relative Position Intensity PositionIntensity Position Intensity (°2θ) (%) (°2θ) (%) (°2θ) (%) 6.6 31 6.6 336.6 24 7.0 42 7.0 35 7.0 21 9.3 16 9.3 24 9.2 19 11.9 100 11.9 100 9.617 13.3 20 13.3 27 11.9 55 14.0 96 14.0 85 13.3 20 14.1 22 14.2 34 13.939 15.1 11 15.1 16 16.8 50 16.8 13 16.8 12 17.1 100 17.1 22 17.1 20 17.756 17.7 14 17.7 15 18.2 38 18.4 69 18.4 65 18.3 37 18.6 13 18.5 17 19.922 20.0 16 20.0 20 20.2 18 20.8 35 20.8 46 20.8 38 21.3 17 21.3 29 21.324 21.7 59 21.7 82 21.7 56 22.0 16 22.0 18 22.5 19 22.9 14 22.9 19 22.917 23.2 9 23.2 16 23.9 26 23.9 17 23.9 17 24.3 29 24.3 12 24.3 14 25.315 26.8 20

Example 11: Stability of Z-Endoxifen Free Base

Commercially, endoxifen is available as an E/Z isomer free base mixture,as well as ≥98% (Z)-endoxifen HCl and (Z)-endoxifen citrate salts.Stability of Z-endoxifen HCl salt in aqueous and solid forms has beenpreviously published (Elkins et al. J Pharm Biomed Anal. 2014 January;88: 174-179). Elkins et al. provide predicted t90 values for 10%conversion of (Z)-endoxifen HCl to (E)-endoxifen in the solid state of53 months at 25° C./60% RH and 4.3 months at 40° C./75% RH. Conversionis expected to be more rapid in a solution environment. In an aqueousmedium, Elkins et al. provide (Z)-endoxifen HCl t90 values of 149 daysat room temperature and 9 days at 45° C. There remains a need for(Z)-endoxifen free base preparations that are sufficiently stable forpreparation of pharmaceutical compositions that are suitable foradministering to subjects, for example, at ambient temperatures as wellas at higher temperature and humidity.

Provided herein in the present disclosure are preparations of endoxifenfree base that are substantially pure, at least 90% (Z)-endoxifen freebase, and stable for at least 9 months under conditions of ambient andhigh temperature and humidity. Stability, for the purpose of thisdisclosure, is defined as the continued presence of at least 90%(Z)-endoxifen in a composition for at least 6 months, and is measurableby (Z)-endoxifen conversion to (E)-endoxifen starting from the date ofsynthesis.

A. Accelerated 10-Day Stability Study of (Z)-Endoxifen Free Base

The stability of (Z)-endoxifen free base Sample 1 of Table 11 wasstudied when stored at various temperatures in solid form and in ethanol(EtOH) solution. The (E)-endoxifen and (Z)-endoxifen levels weremeasured on days 2, 7 and 10 following placement of 1 g aliquots ofpurified (Z)-endoxifen free base in 180 mL HDPE bottles under dry N₂,which were then placed in double LDPE bags. The double bagged HDPEbottles containing each sample were individually packed in aluminum foilbags and thermally sealed until they were opened for testing thestability of the samples. The (E)- and (Z)-isomers of endoxifen weremonitored by HPLC as disclosed above. The purity of (Z)-endoxifen insolid form remained at about 97% at 40° C., 60° C., and 80° C. up to 10days. The results of the accelerated 10 days study indicate that(Z)-endoxifen in solid form does not interconvert to (E)-endoxifen atthe tested temperatures up to 10 days.

TABLE 11 Accelerated Stability Test of Solid (Z)-endoxifen Free Base No.1 2 3 Z-endoxifen 1 g 1 g 1 g Temperature 40° C. 60° C. 80° C. 2 E/Z1/76 (1.27%/97.45%) 1/75 (1.30%/97.43%) 1/76 (1.28%/97.42%) days ratioTotal 1.28% 1.27% 1.30% Impurity 7 E/Z 1/81 (1.2%/97.11%) 1/83(1.17%/87.11%) 1/7 2(1.34%/96.92%) days ratio Total 1.69% 1.72% 1.74%Impurity 10 E/Z 1/75 (1.29%/97.31%) 1/82 (1.27%/97.48%) 1/77(1.27%/97.33%) days ratio Total 1.40% 1.33% 1.40% Impurity

TABLE 12 Accelerated Stability Study of (Z)-endoxifen Free Base Solutionin Ethanol No. 1 2 3 4 (Z)-endoxifen 0.1 g 0.1 g 0.05 0.05 g Solvent 10mL 10 mL 18 mL 18 mL 99.5% EtOH 99.5% EtOH 70% EtOH 70% EtOHConcentration 10 mg/mL 10 mg/mL 2.8 mg/mL 2.8 mg/mL Temperature 25° C.40° C. 25C° C. 40° C. 2 E/Z 1/71 1/57 1/46 1/21 days Ratio(1.37%/97.33%) (1.70%/96.98%) (2.09%/96.59%) (4.43%/94.23%) Total 13%1.32% 1.33% 1.34% Impurity 7 E/Z 1/50 1/34 1/17 1/5.5 days Ratio(1.91%/96.37%) (2.82%/95.45%) (5.62%92.67%) (15.23%/83.04%) Total 1.72%1/73% 1.71% 1.73% Impurity 10 E/Z 1/37 1/27 1/8.4 1/3.4 days Ratio(2.58%/96.03%) (3.56%/95.10%) (10.46%/88.20%) (22.59%/76.09%) TotalImpurity 1.39% 1.34% 1.34% 1.32%

Results show that (Z)-endoxifen free base is surprisingly stable inalcoholic (for example ethanol and isopropanol (data not shown))solutions even at elevated temperatures (40° C.) at higherconcentrations over 10 days. Accelerated stability at highertemperatures are generally considered predictive of long term (at least18 m) stability at ambient temperatures.

B. Bulk Drug Stability Study

(Z)-endoxifen free base of the instant disclosure prepared by methodsdisclosed herein is surprisingly stable in bulk form. For bulk stabilitystudies, ˜1 g aliquots of (Z)-endoxifen free base were placed insideindividually in double LDPE bags tied with a nylon cable tie. The LDPEbags were then placed in aluminum foil bags and thermally sealed. Thesamples for stability were placed in tightly capped, 180 mL HDPE bottlesunder inert conditions (dry nitrogen) at 5° C. and 25° C./60% RH for 12months, and at 40° C./75% RH for 3 months. The samples were tested for(E)-endoxifen and (Z)-endoxifen concentrations and impurities, moistureor water content by Karl Fischer titration, aerobic bacterial colonyforming units and appearance at time points day 0, 10 days, 1 m and 3 m.Table 13 below provides bulk drug stability data at 9 m on endoxifenfree base at varying storage conditions.

TABLE 13 Bulk Drug Stability of (Z)-endoxifen Free Base % Purity %Purity % Purity Storage Storage Endoxifen Storage (5° C.) 25° C./60% RH40° C./75% RH (Free Base) 0 m 3 m 6 m 9 m 0 m 3 m 6 m 9 m 0 m 3 m(Z)-endoxifen 99.5 99.6 99.5 99.5 99.5 99.6 99.5 99.5 99.5 99.5(E)-endoxifen 0.19 0.17 0.18 0.19 0.19 0.17 0.18 0.19 0.19 0.17 Watercontent 0.06 0.05 0.05 0.05 0.06 0.05 0.06 0.07 0.06 0.07 (Karl Fischer)Max Indiv. 0.11 0.10 0.11 0.11 0.11 0.11 0.11 0.11 0.11 0.11 Impurity(Z)-endoxifen 13990 14030 13776 USP < 921 ≥ Plate Count (cfu g/mL)

Results show that solid Z-endoxifen free base is stable for at least 9months at 5° C. and 25° C./60% RH. Accelerated stability at hightemperatures is considered to be predictive of long term (at least 18 m)stability at ambient temperatures. (Z)-to-(E) endoxifen interconversionis minimal at the storage conditions studied, water content was not morethan 1% and aerobic bacterial plate count TAC (cfu) is less than 20,000cfu g/mL. Maximum individual impurities were determined at each timepoint and ranged from 0.10% to 0.11%.

Example 12: Characteristics of Purified Solid Z-Endoxifen Free Base

(Z)-endoxifen free base, produced by methods disclosed herein, was awhite to off-white powder. The water content of such a powder was notmore than 1% as determined using Method Ic (Karl Fischer titration) ofUSP 921. The residual solvents were measured. Methanol was not more than(NMT) 3000 ppm, tetrahydrofuran NMT 720 ppm, isopropanol, ethyl acetate,n-heptane, and ethanol were each NMT 5000 ppm.

Residue on ignition of (Z)-endoxifen was not more than 0.1% asdetermined by Method II of USP 281. Residue on ignition of (Z)-endoxifenranged from 0.02% to 0.099%. In some embodiments, the residue uponignition is not more than 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%,0.08%, 0.09%, and 0.1%. In another aspect, the heavy metals were NMT 20ppm as tested by Method II of USP 231.

Elemental levels in solid (Z)-endoxifen free base is tested using testmethods of USP 232 and USP 40.

Thus, the processes described herein and endoxifen prepared using theseprocesses are suitable for manufacture of clinical and commercial grade(Z)-endoxifen.

FTIR Spectrum of Z-Endoxifen

(Z)-endoxifen was analyzed using Fourier Transform Infra-RedSpectroscopy (Instrument type: FT/IR4600Type A; Jasco: S/N ratio25,000:1, Max resolution 0.7 cm⁻¹) with standard light source and a TGSdetector. Infrared spectra were recorded on a FTIR spectrometer (ModelFT/IR 4600 Type A; manufacturer Jasco), equipped with triglycine sulfate(TGS) detector and KBr beam splitter, using AgBr windows. About 10 mg of(Z)-Endoxifen free base was placed onto the ATR crystal, pressure wasapplied with the tip, and the sample spectrum was collected. Spectrawere collected with a scan speed to 2 mm/sec using hydrated films.Interferograms were accumulated over the spectral range 4000-650 cm⁻¹with a nominal resolution of 0.4 cm⁻¹ and 32 scans. FIG. 2 shows arepresentative IR Spectrum of (Z)-endoxifen free base.

FIG. 2 shows that purified (Z)-endoxifen has peaks at 702.0, 730.9,794.5, 825.4, 907.3, 1027.9, 1167.7, 1238.1, 1278.6, 1508.1, 1575.6,1607.4, 3310.2 wavelengths cm⁻¹+/−0.4 cm⁻¹. A previously published FT/IRspectrum of free endoxifen showed peaks are at 706, 831, 1053, 1171,1240, 1465, 1507 and 1604 wavelength cm⁻¹ (Agudelo et al. PLoS ONE 8(3):e60250. doi:10.1371/journal.pone.0060250).

TABLE 14 Spectral Assignments for IR spectrum of (Z)-endoxifen free baseAssignment Frequency Strength N—H stretching 3200-3500 weak O—Hstretching C═C stretching 1500 1680 Medium CH₂ & CH₃ bending 1200-1500Medium C—N stretching 1000-1200 Medium C—O stretching ═C—H bending 690-1000 Medium ═CH₂ bending

Example 13: Oral Compositions of Z-Endoxifen

API-in-Capsule.

≥90% Z-endoxifen free base was prepared as disclosed above. Drywhite-to-off-white powder is formulated as stable free-flowing powderand filled neat into a capsule as a Drug-in-Capsule (DIC, also calledAPI-in-capsule, AIC) oral solid dosage form. 1 mg, 2 mg, and 4 mg ofendoxifen which was at least 90% (Z)-endoxifen was filled neat in aVCaps® Plus enteric capsule using Xcellodose technology (Capsugel). AICwere of size 0, Swedish orange in color.

The VCaps® Plus enteric capsules (Capsugel) are made with gluten free,non-animal self-gelling product hypromellose (methyl and hydroxypropylmixed ether of cellulose), with low-moisture content that is suitablefor moisture-sensitive ingredients such as (Z)-endoxifen. The capsulesare coated with an enteric coating designed to achieve intestinaltargeting (upper GI and colon), by the method of Cole et al. (Cole etal., Int. Journal of Pharmaceutics Vol. 231 83-95, 2002). Eudragit FSD30 is used as enteric coating for colonic targeted release and EudragitL30 D55 is used as enteric coating for upper gastrointestinal targetedrelease.

(Z)-endoxifen free base in the AIC is released in predominantly in theintestines (upper GI and colon) and is protected from the acidicenvironment in the stomach for at least 6 hours. Enteric coating of thecapsule prevents the release of (Z)-endoxifen in the stomach for atleast 6 hours as tested by a method of USP 711.

Enteric AIC disclosed herein is stable for at least 6 months.

Standards and Testing for Microbial Contamination of the Oral SolidDosage Forms of the Instant Disclosure.

For formulations of the solid dosage compositions disclosed herein, asthe water content is NMT 1% and water activity (Aw) is less than 0.75,testing TAC and USP indicator organism is not necessary. Thepublication, “Microbial Bioburden on Oral Solid Dosage Form,” by Jose E.Martinez, Pharmaceutical Technology, February 2002, pages 58 to 70, ishereby incorporated by reference in its entirety.

Furthermore, since formulations of the compositions disclosed hereinalso have water activity of less than 0.75, then no detailed microbialtesting of that need be done. Total Aerobic Plate Count (TAC) is anestimation of the total viable aerobic bacteria present in a sample ofraw material, in-process material, or finished product. Samples areanalyzed in accordance with the most current USP Guidelines Chapter 61(Microbial Limits Tests).

Acceptable TAC for oral solid dosage forms (OSDFs) are established forthe formulation of the inventive compositions in terms of alert andaction levels, which could be 1000 cfu g/ml, and 10,000 cfu g/mLrespectively. A TAC that is 20,000 cfu g/mL is considered unacceptable.

Example 14: Preparation of (Z)-Endoxifen Salt

(Z)-Endoxifen D-gluconate salt is prepared by mixing an ethanolic slurryof (Z)-endoxifen as the free base with an aqueous solution of D-gluconicacid, followed by hydrolyzing a 20% w/v solution of D-gluconolactone inwater and heating at 70° C. for 15 to 30 min. Minimum volume of ethanolis used and 5 ml of the aqueous D-gluconic acid solution is adder per 1g of endoxifen free base. Stirring is then continued until a clearsolution is obtained. (Z)-Endoxifen D-gluconate is crystallized usingthe methods disclosed above in Examples 1 to 4 to provide (Z)-endoxifenD-gluconate salt.

1 mg, 2 mg, 5 mg, 10 mg, 20 mg and 40 mg AIC enteric capsules will beprepared as described above. Similarly, 1 mg to 50 mg tablets will beprepared using (Z)-endoxifen D-gluconate salt with enteric coatingsEUDRAGIT® FS D30 and EUDRAGIT L30D 55 to target small intestine andcolon.

TABLE 15 Exemplary Tablet wt/wt of Ingredient composition (Z)-endoxifenD-gluconate 0.1% to 20% croscarmellose sodium 0.5% to 15%microscrystalline cellulose 15% to 90% mannitol 5% to 30% Entericcoating 0.01% to 3% EUDRAGIT ® L30D 55

Example 15: Placebo Controlled, Dose Escalation Safety andPharmacokinetic Study of Oral (Z)-Endoxifen

An objective of the study was to assess the safety and tolerability oforal (Z)-Endoxifen when administered as a topical application to thebreast or orally as a capsule to healthy female volunteers. A secondaryobjective was to assess the pharmacokinetics of multiple doses of oral(Z)-Endoxifen in the subjects.

Generally, healthy female volunteers aged ≥18 years and ≤65 years wereenrolled. 24 participants were enrolled in 3 cohorts and will beadministered study drug orally. Three dose levels of orally administered(Z)-Endoxifen were investigated in 3 cohorts. In each cohortparticipants were randomized to receive an oral (Z)-Endoxifen or placeboin a blinded fashion. Enteric-resistant capsules used in the study weredesigned as delayed release capsules to resist rupture for at least 30minutes, but typically, for about 2 hours, under gastric conditions andthen open fully in intestinal fluid (DRCaps; Capsugel, a Lonza Company,USA). Enteric-resistant (Swedish Orange size Zero (0); Capsugel, USA)DRCaps capsules manually filled with 1, 2 or 4 mg of (Z)-endoxifen neatin compliance with GMP requirements were used. Placebo capsulescontained microcrystalline cellulose an inert common capsule and tabletexcipient.

Each cohort enrolled 8 participants, with 6 participants receiving(Z)-Endoxifen 1 mg capsules orally at 1 mg (Cohort 1), 2 mg (Cohort 2)or 4 mg (Cohort 3) doses and 2 participants receiving matching placebocapsules. In Cohort 1 (Dose Level=1 mg) two participants (sentinels)were dosed 24 hours prior to the remaining participants. One sentinelwas dosed with (Z)-Endoxifen and the other with placebo. The remainingsix participants in Cohort 1 were dosed as no safety concerns wereidentified in the sentinel participants. The non-sentinel participantsin Cohort 1 were admitted to the clinical research facility at least oneday later than the sentinels. There were no sentinels in Cohorts 2 and3.

Participant engagement in this study was for a period of 56 days. Theseincluded a 28 day screening period, and a 7 day post treatment periodfor Pharmacokinetic (PK) sampling. There were two confinement periods,one at the beginning and one at the end of the dosing periods.Participants were confined to the clinical facility on Days −1 throughto Day 2 and Days 21 and 22. If participants were to experience anyclinically significant adverse events during the study, they would havebeen required to remain in the clinical facility for further observationat the discretion of the Principal Investigator (PI).

Healthy female volunteers were screened within 28 days prior tocommencement of dosing. Participants were admitted to the clinicalfacility on Day −1 for up to 3 days. The first dose of (Z)-Endoxifen orplacebo capsules was administered on Day 1. Following completion of allsafety assessments and sampling for PK analyses, participants weredischarged from the clinical facility on Day 2. The first dose on Day 1was followed by a 6-day treatment free period (Days 2-7). Eachparticipant returned to the clinical facility for PK blood draws andsafety assessments during the treatment free period on study Days 4 and6. On Day 8 participants commenced daily administration of (Z)-Endoxifencapsules or placebo capsules for 14 consecutive days (Days 8-21).Participants were supplied with study drug capsules and asked to selfadminister daily. Participants visited the clinical facility on Days 11,14 and 17, prior to dose administration, for PK blood draws and safetyassessments. On the morning of Day 21 each participant returned to theclinical facility prior to dose administration and were confined to thefacility until Day 22 to allow for the collection of PK blood draws andsafety assessments. The Day 21 dose was the last administration of studydrug. Participants were discharged from the clinical facility on Day 22and returned for PK blood draws and safety assessments on study Days 24,26, and 28.

Study assessments included taking the subjects medical history,including evaluation of any on-study adverse events and concomitantmedication use; height and weight; physical examination; periodic vitalsigns (body temperature, heart rate, respiratory rate, blood pressure);periodic 12-lead ECGs. Laboratory tests included hematology,coagulation, urinalysis, serum chemistry and biomarker analysis (e.g.,CYP2D6, BRCA1/2, Ki67, tamoxifen metabolites, etc.). Specificassessments to evaluate treatment safety included the following: thefrequency and type of adverse events, clinical laboratory testing,12-lead ECGs and vital signs. A modified FACT-ES® scoring questionnairewas used to assess symptomatology.

Blood draws for PK analysis were collected pre-dose (within 10 min) and0.5, 1, 2, 3, 4, 6, 8, 10, 12 and 24 hours following study drugadministration on Days 1 and 21. PK samples for all other study dayswere collected within 10 min prior to dose administration. PK sampleswere obtained during the treatment free period on Days 4 and 6.Additional, PK samples were collected during the daily dosing period onDays 8, 11, 14 and 17 (within 10 min pre-dose). PK samples followingcompletion of treatment will be collected on Days 24, 26 and 28.AUC(inf) was measured on Day 1 blood draw samples. Steady state plasmalevels were determined starting from Day 8 onwards (which served as Day0) for this determination.

Safety and Tolerability

Safety endpoints were summarized by dose cohort, with placebos pooledacross cohorts. Treatment-emergent AEs were coded using the latestversion of MedDRA by System Organ Class (SOC) and Preferred Term,classified from verbatim terms. The incidence and frequency of AEs, andSAEs, were summarized by cohort according to SOC and Preferred Terms,and by severity and relationship. The duration of AEs was determined andincluded in listings, along with the action taken and outcome. Vitalsigns, ECG and safety laboratory parameters were summarized at eachscheduled time point using descriptive statistics. Post-dose assessmentswere compared with baseline measurements. The incidence of laboratoryabnormalities was summarized. Physical examination findings werepresented in listings.

Treatment-emergent AEs deemed to be related to study drug (probably orpossibly) were reported in 15 of 18 subjects (83%, 41 AEs) who received(Z)-Endoxifen and in 4 of 6 subjects (67%, 20 AEs) who received placebo.Most treatment-related AEs were mild in severity, with treatment relatedAEs of moderate severity reported in 5 of 18 subjects (28%, 6 AEs) whoreceived (Z)-Endoxifen and in 2 of 6 subjects (33%, 2 AEs) who receivedplacebo.

All AEs were classified as mild (75 of 84 AEs) or moderate (9 of 84 AEs)in severity, with no AEs classified as severe. Common adverse events oftension headache, headache, abdominal pain, nausea, dysmenorrhea andfatigue were reported both in subjects who received oral (Z)-Endoxifenand subjects who received placebo. Upper respiratory infection, hotflush, abdominal distension, dry mouth, and menstruation delayed werereported only in subjects who received (Z)-Endoxifen.

There were no clear differences in safety as assessed by clinicallaboratory tests, vital signs, ECG assessments, FACT-ES responses insubjects who had received either topical or oral (Z)-Endoxifen comparedwith those who received placebo, and no dose-related trends in thesesafety assessments in those subjects who received (Z)-Endoxifen.

Pharmacokinetics

Mean and individual (Z)-Endoxifen serum concentration-time curves weretabulated for each dose cohort, and presented graphically withconcentration displayed on a linear and logarithmic scale.Pharmacokinetic parameters were determined for each participant andsummarized by cohort using descriptive statistics (arithmetic means,standard deviations, coefficients of variation, sample size, minimum,maximum and median). In addition, geometric means was calculated for AUCand Cmax. Analyses using linear models was performed to assess doseproportionality (both after a single dose and multiple dose), timedependence and accumulation (oral multiple dose), and attainment ofsteady state (multiple dose).

Parameters determined for the first and last dose (Days 1 and 21)included time to maximum concentration (Tmax), maximum concentration(Cmax), area under the concentration-time curve from time 0 to 24 hoursfollowing drug administration (AUC_(0-24 h)), terminal elimination rateconstant (kel), terminal half-life (t_(1/2)), terminal clearance (CL/F)and volume of distribution (Vd/F). Area under the concentration-timecurve from time 0 to infinity (AUC_(0-inf)) was also determined for thefirst dose, on Day 1.

The pharmacokinetic parameters were determined using non-compartmentalmethod(s). Descriptive statistics of pharmacokinetic parameters includedmean, standard deviation (SD), and coefficient of variation (CV),minimum (min) and maximum (max). Dose related trends in pharmacokineticparameters were assessed.

Mean and individual endoxifen serum concentration-time curves weretabulated for each dose cohort. Pharmacokinetic parameters, weredetermined for each participant and summarized by cohort usingdescriptive statistics (arithmetic means, SD, CV, sample size [N], min,max and median). In addition, geometric means were calculated for AUCand Cmax. Analyses using linear models were performed to assess doseproportionality (both after a single dose (oral) and multiple dose(topical and oral)). Statistical analysis was performed on thepharmacokinetic parameters using SAS v9.3 and Phoenix WinNonLin version7.0 or higher.

Abbreviations Used:

CV Coefficient of Variance

Tmax time to maximum concentration

Cmax maximum concentration,

AUC_(0-24 h) (“AUC24 hr”) area under the concentration-time curve fromtime 0 to 24 hours following drug administration

kel terminal elimination rate constant, and volume of

t1/2 terminal half-life

CL/F terminal clearance

Vd/F distribution.

AUC_(0-inf) (“AUC_(0-inf)”) Area under the concentration-time curve fromtime 0 to infinity was also determined for the first dose, on Day 1.

A summary of serum pharmacokinetic parameters with terminal half-life of(Z)-endoxifen, terminal elimination half-life, Tmax, Cmax, and AUC areprovided below in Table 16.

TABLE 16 Model Serum Pharmacokinetic Parameters (Z)- Endoxifen MedianArithmetic Mean (CV %) Dose/Day (Range) C_(max) AUC_(24hr) t_(1/2) Day(mg) T_(max) (hr) (ng/mL) (hr*ng/mL) (hr) Day 1 1 mg 6.0 9.6 158 41.1 (n= 5) (2.0-8.08) (18.9%) (20.3%) (15.8%) 2 mg 4.17 17.7 305 49.0 (n = 6)(3.0-6.0)  (23.4%) (17.9%) (26.0%) 4 mg 5.08 32.5 587 41.6 (n = 6)(3.0-6.0)  (52.3%) (53.3%) (22.9%) Day 21 1 mg 8.0 24.6 458 52.6 (n = 5)(4.0-6.05) (35.0%) (36.3%) (20.8%) 2 mg 4.0 50.0 911 43.4 (n = 6)(2.0-6.05) (23.7%) (19.7%) (17.1%) 4 mg 5.0 115.0 2041 45.1 (n = 6)(2.0-10.0) (49.0%) (50.4%) (15.2%)

For once daily oral administration of the delayed-release(acid-resistant) (Z)-endoxifen capsules, there was dose-proportionality(linear dose response) in Cmax and AUC_(24 h) of endoxifen across thethree dose levels used in the study. Cmax increased from 9.6 ng/mL to32.5 ng/mL for the 1 mg to 4 mg doses on day 1 and from 24.6 ng/mL to115.0 ng/mL on Day 21. Median Tmax by dose level on Days 1 and 21 rangedfrom 4 to 8 hours. The apparent terminal half life (t_(1/2)) by doselevel ranged between 42 and 53 hours, and steady state appeared to beattained after approximately 7 days (FIG. 5—dosing at Day 8 and Css atDay 14). Time to reach maximum blood levels ranged from 4 to 8 hours(FIG. 4).

TABLE 17 Model AUC_((0-inf)): 1 mg. 2 mg, and 4 mg (Z)-endoxifenAUC_((0-inf)) hr* ng/mL Dose: 1 mg Dose: 2 mg Dose: 4 mg 441 755 736 534954 1883 739 1242 2116 527 1489 4624 593 1009 1098 1188 1642

TABLE 18A Model Serum Concentrations (nM) Over Time by Treatment, Dose:1 mg (Z)-Endoxifen Day 8 Day 11 Day 14 Day 17 Pre- Day 1 Day 4 Day 6Pre- Pre- Pre- Pre- Statistic dose hr 0.5 hr 1 hr 2 hr 3 hr 4 hr 5 hr 8hr 10 hr 12 hr 24 Hr 72 Hr 120 dose Dose dose Dose Mean 2.68 2.68 3.7517.1 17.1 20.3 23.6 22.0 22.5 18.2 15.0 8.03 3.75 2.68 38.5 43.4 43.4Std Dev 0.0 1.89 2.39 2.4 1.5 3.6 4.4 6.9 6.7 5.8 3.1 1.89 1.47 0.00 8.622.8 19.1 CV (%) 0 71 64 14 9 18 19 32 30 32 20 24 39 0 22 53 44 Median2.68 2.68 2.68 18.7 16.1 18.7 24.1 21.4 21.4 21.4 16.1 8.03 2.68 2.6842.8 40.2 45.5 Min 2.68 0.00 2.68 13.4 16.1 18.7 16.1 13.4 13.4 8.0310.7 5.35 2.68 2.68 26.8 13.4 13.4 Max 2.68 5.35 8.03 18.7 18.7 26.826.8 32.1 32.1 21.4 18.7 10.7 5.35 2.68 45.5 75.0 64.2 GeoMean 2.68 3.183.33 17.0 17.1 20.1 23.2 21.1 21.6 17.1 14.7 7.84 3.53 2.68 37.7 37.638.5

TABLE 18B Model Pharmacokinetic Parameters by Treatment; Dose 2 mg(Z)-Endoxifen (n = 6). Mean 0.17 0.00 2.17 10.2 14.5 17.2 17.2 14.3 14.514.2 10.3 5.67 2.80 0.83 22.8 29.2 32.2 Std Dev 0.41 0.00 1.47 3.8 2.34.8 3.7 3.8 3.4 2.0 2.3 1.53 1.10 0.43 3.3 4.7 7.7 CV (%) 245% 0% 58%37% 16% 28% 23% 26% 23% 34% 22% 29% 39% 49% 15% 16% 24% Median 0.00 0.002.50 10.0 14.5 17.5 17.0 14.5 14.5 14.0 10.5 5.50 3.00 1.00 22.5 28.529.5 Minimum 0.00 0.00 0.00 5.00 11.0 11.0 13.0 8.00 10.0 12.0 8.0 4.001.00 6.00 19.0 23.0 27.0 Maximum 1.00 0.00 4.00 16.0 17.0 22.9 22.0 19.019.0 18.0 10.0 8.00 4.00 1.00 28.0 37.0 47.0 GeoMean 1.00 2.35 8.55 14.316.6 16.8 13.8 14.2 14.1 10.1 5.47 2.55 1.00 22.5 28.9 31.5

TABLE 18C Model Pharmacokinetic Parameters by Treatment; Dose 4 mg(Z)-Endoxifen (n = 6) N 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 Mean 0.83 0.833.17 20.0 27.8 26.8 30.5 25.5 30.3 27.8 21.3 10.0 5.50 2.33 50.4 69.088.8 Std Dev 0.41 0.41 1.72 5.0 13.7 12.1 18.0 11.0 17.8 14.9 13.2 6.84.54 1.97 25.3 45.8 43.0 CV (%) 49% 49% 54% 40% 49% 45% 59% 43% 59% 53%52% 68% 93% 84% 50% 86% 62% Median 1.00 1.00 4.00 19.0 24.5 24.0 25.525.0 28.0 25.5 19.5 8.00 4.00 3.50 45.0 64.0 73.0 Minimum 0.00 0.00 1.0011.9 14.0 14.0 14.0 13.0 18.0 11.0 8.00 4.00 3.00 1.00 33.0 3.00 1.00Maximum 1.00 1.00 5.00 35.0 52.0 49.0 62.0 42.0 62.0 52.0 43.0 23.0 10.06.00 77.0 145 115 GeoMean 1.00 1.00 2.52 38.9 25.3 24.8 26.7 23.5 35.424.6 18.0 8.54 3.82 1.92 45.2 44.9 37.5

TABLE 19A Day 1 Model Pharmacokinetic Parameters by Treatment; Dose 1 mg(Z)-Endoxifen. N = 5 Day 1 Cmax Tmax AUCTlast Kel Thalf AUC_(0-inf)AUC24hr CL/F Vz/F Statistic (ng/mL) (hr) (hr * ng/mL) (l/hr) (hr) (hr *ng/mL) (hr * ng/mL) (L/hr) (L) Mean 9.6 5.62 471 0.015 46.1 567 158 1.82119.9 Std Dev 1.8 2.21 76 0.002 7.3 110 32 0.33 27.4 CV (%) 18.9 39.416.2 16.2 15.8 19.10 20.8 18.4 22.9 Median 10.0 6.0 472 0.015 47.2 534171 1.87 107.6 Min 7.0 2.0 367 0.013 37.5 441 102 1.35 102.7 Max 12.08.08 582 0.018 54.4 739 183 2.27 168.4 GeoMean 9.5 5.11 466 0.015 45.6559 155 1.79 117.8 GeoCV % 19.9 58.2 16.5 16.2 16.2 19.1 24.0 19.1 20.6

TABLE 19B Day1 Model Pharmacokinetic Parameters by Treatment; Dose 2 mgN 6 6 6 5 6 6 6 6 6 Mean 17.7 4.28 889 0.015 49.0 1105 385 1.89 130.5Std. Dev 4.1 0.96 225 0.003 12.7 256 55 0.46 34.9 CV (%) 23.4 22.5 25.323.3 26.0 23.3 17.9 24.1 25.7 Median 17.5 4.17 955 0.015 44.0 1099 3031.83 118.0 Minimum 13.0 3.00 537 0.010 37.8 755 248 1.34 98.2 Maximum22.2 6.00 1137 0.018 59.1 1499 393 2.55 194.5 GeoMean 17.3 4.19 8620.015 47.7 1081 301 1.85 127.3 GeoCV % 24.1 22.2 28.7 25.3 25.3 24.018.0 24.0 24.7

TABLE 19C Day 1 Model Pharmacokinetic Parameters by Treatment; Dose 4 mgN 6 6 6 6 6 6 6 6 6 Mean 32.5 5.56 1654 0.017 41.6 2017 587 2.73 154.5Std. Dev 17.0 2.47 1047 0.004 9.5 1375 313 1.60 79.9 CV (%) 52.3 44.563.3 21.8 22.9 58.2 53.3 58.5 51.7 Median 29.5 5.08 1423 0.017 39.9 1763539 2.28 130.7 Minimum 14.0 3.00 691 0.012 31.6 736 258 0.87 69.9Maximum 62.0 10.00 2574 0.022 56.0 4524 1116 5.43 259.4 GeoMean 29.15.15 1424 0.017 40.7 1704 521 2.35 137.8 GeoCV % 55.7 43.6 64.4 22.822.8 59.1 57.8 59.1 56.5

TABLE 19D Day 21 Model Pharmacokinetic Parameters by Treatment; Dose 1mg (Z)-Endoxifen. N = 5 N 5 5 5 5 5 5 5 5 Mean 24.6 6.98 458 19.1 0.01452.6 2.49 181.3 Std. Dev 8.6 1.87 166 6.9 0.003 10.9 1.06 64.1 CV (%)35.0 27.1 38.3 36.3 20.4 20.8 42.6 35.4 Median 29.0 8.00 527 22.0 0.01449.3 1.90 170.1 Minimum 14.0 4.00 248 10.3 0.011 40.6 1.57 116.7 Maximum34.0 8.42 635 26.5 0.017 65.8 4.03 286.8 GeoMean 23.3 6.64 431 17.90.013 51.7 2.32 173.3 GeoCV % 40.1 32.1 42.2 42.2 20.9 20.9 42.2 33.9

TABLE 19E Day 21 Model Pharmacokinetic Parameters by Treatment; Dose 2mg (Z)-Endoxifen. N 6 6 6 6 6 6 6 6 Mean 50.0 4.01 911 38.0 0.016 43.42.27 139.1 Std Dev 11.9 1.80 179 7.5 0.003 7.4 0.44 22.7 CV (%) 23.744.9 19.7 19.7 18.9 17.1 19.4 16.3 Median 51.0 4.00 910 37.9 0.015 45.72.21 131.5 Minimum 35.0 2.00 716 29.8 0.014 33.0 1.70 119.6 Maximum 65.06.05 1177 49.1 0.021 50.8 2.79 179.4 GeoMean 48.8 3.64 897 37.4 0.01642.8 2.23 197.7 GeoCV % 24.9 53.1 19.8 19.8 18.2 18.2 19.8 15.5

TABLE 19F Day 21 Model Pharmacokinetic Parameters by Treatment; Dose 4mg (Z)-Endoxifen. N 6 6 6 6 6 6 6 6 Mean 115.0 5.33 2041 85.1 0.016 45.12.83 189.0 Std Dev 56.3 2.73 1029 42.9 0.003 6.8 2.15 156.0 CV (%) 49.051.2 50.4 50.4 16.6 15.2 76.0 82.5 Median 131.0 5.00 2469 102.9 0.01545.3 1.63 106.1 Minimum 42.0 2.00 616 25.7 0.013 34.0 1.32 73.0 Maximum178.0 10.00 3024 126.4 0.020 54.3 6.49 461.5 GeoMean 100.5 4.75 174472.7 0.016 44.6 2.29 147.7 GeoCV % 67.4 58.5 76.1 76.1 15.9 15.9 76.184.6

TABLE 20 Mean Ratios of Pharmacokinetic Parameters (Day21)/(Day1) MeanRatios of Pharmacokinetic (Z)- Parameters (Day 21)/(Day1) EndoxifenC_(max)(Day21)/ AUC_(24hr)(Day21)/ AUC_(24hr)(Day21)/ Dose C_(max)(Day1)AUC_(24hr)(Day1) AUC_(0-inf)(Day1) 1 mg (n = 5) 2.74 (SD 1.29) 3.09 (SD1.49) 0.81 (SD 0.20) 2 mg (n = 6) 2.92 (SD 0.88) 3.04 (SD 0.79) 0.85 (SD0.20) 4 mg (n = 6) 3.56 (SD 1.01) 3.47 (SD 1.08) 1.08 (SD 0.38)

The mean accumulation ratios ranged from 2.74 to 3.56 for Cmax and from3.04 to 3.47 for AUC_(24 h) as the ratio of parameters on Day 21compared with Day 1. There was no marked trend by dose level inclearance (Cl/F) and volume of distribution (Vd/F) for Day 1 and Day 21.

Time to maximum serum endoxifen levels is achieved by about 10 hours andthat there is dose-related increase in serum endoxifen levels (FIG. 3and FIG. 4). Serum endoxifen levels ranged from about 25 nM to about 80nM.

Pharmacokinetic assessment of pre-dose trough concentration from Day 8to Day 21 (FIG. 4) showed attainment of straight line after day 14.Steady state levels of serum endoxifen appear to be being achieved byDay 7 with daily dosing. The pre-dose serum concentrations indicatelittle accumulation after 17 days for 4 mg of (Z)-Endoxifen.

TABLE 21 Parameter estimates of dose linearity for endoxifenPharmacokinetic β Two-sided 95% Parameter (log) Analyte Day EstimateConfidence Interval r² Cmax Endoxifen 1 0.808 (0.487-1.130) 0.66 211.055 (0.655-1.455) 0.68 AUC_(24hr) Endoxifen 1 0.873 (0.548-1.199) 0.6921 1.008 (0.582-1.433) 0.63

TABLE 22 Comparison of (Z)-endoxifen concentrations (Oral) withpublished literature. Published Literature % Result of Study Result ofRatio (Z)- Average Plasma Average Serum of Css Endoxifen (Z)-EndoxifenCss (Z)-Endoxifen Css Study/ Dose ng/mL nM ng/mL nM Literature 1 mg 12.433.1 19.1 51.1 154% 2 mg 18.6 49.8 38 101.7 204% 4 mg 56.8 152.1 85.1227.9 150% 8 mg 96.8 259.2 — — — Css = Average Concentration at SteadyState (AUCtau/tau)

PUBLISHED LITERATURE

-   1. Wu X et al. Cancer Res. 2009 Mar. 1; 69(5):1722-7. PubMed PMID:    19244106-   2. Ahmad A et al. Breast Cancer Res Treat. 2010 July; 122(2):579-84.    PubMed PMID:20052538-   3. Ahmad A et al. J Clin Oncol 30, 2012 (suppl; abstr 3089)-   4. Goetz M P et al. San Antonio Breast Conference 2013 [PD3-4]-   5. Goetz M P et al. San Antonio Breast Conference 2015. [PD203]-   6. Ahmad A et al. Clin Transl Sci. 2016 Jun. 27.    doi:10.1111/cts.12407. PubMed PMID: 27346789-   7. Lee O et al. Cancer Chemother Pharmacol. 2015 December;    76(6):1235-46

Thus, data from this study suggest that safety and tolerability of oralendoxifen is generally comparable to oral tamoxifen and that subjectsadministered with (Z)-endoxifen compositions have plasma endoxifenlevels that are within the range that has been shown to have therapeuticeffect in the adjuvant setting in women with breast cancer.(Z)-Endoxifen is rapidly absorbed and systemically available and that itdisplays dose proportionality in peak drug concentrations in serum(C(max)) and area under the concentration-time curve extrapolated from 0to infinity (AUC_(0-inf)) over the dose range 1 mg-4.0 mg.

Example 16: Therapeutic Treatment of Tamoxifen-Refractory Breast Cancerby Oral Administration of Z-Endoxifen Free Base

An objective of the study is to demonstrate that stable and therapeutic(Z)-endoxifen levels can be achieved in tamoxifen-refractory patients bysupplementing tamoxifen with (Z)-endoxifen. Plasma endoxifen levels willbe used as a surrogate endpoint to predict clinical benefit as well asthe rate of recurrence in this population and to confirm the safety andtolerability of (Z)-endoxifen administration when compared withtamoxifen.

Generally healthy subjects with early stage estrogen receptor positivebreast cancer who are on adjuvant therapy of tamoxifen for at least 30days will be enrolled for the study for up to 6 months. At leastseventy-five tamoxifen-refractory breast cancer subjects will beenrolled in the study to ensure an evaluable population consists of atleast 25 tamoxifen and 25 (Z)-endoxifen subjects is achieved. Additionalsubjects may be enrolled to achieve the desired population oftamoxifen-refractory subjects. These subjects will have been diagnosedwith breast cancer and undergone mastectomy or lumpectomy.

After at least 30 days of oral tamoxifen, endoxifen plasma levels willbe measured. If the endoxifen level is below 30 or 40 nM, then 1 mg oforal (Z)-endoxifen will be added to the oral tamoxifen adjuvant regimen.Additional (Z)-endoxifen doses will be added until a stable endoxifen ofat least 40 nM but not more than 80 nM is achieved. The endpoint is toestablish a therapeutically stable endoxifen level in thetamoxifen+endoxifen group for at least 6 months.

Example 17: Pre-Surgical Treatment of Estrogen Receptor Positive BreastCancer by Oral Administration of Z-Endoxifen-D-Gluconate

An objective is to determine if Z-Endoxifen-D-gluconate reduces tumoractivity in pre-surgical estrogen receptor positive breast cancerpatients. Upon the initial diagnosis of ER+ breast cancer, 8 patientswill be assigned to one of 3 groups, where each patient will receive 1,2, or 4 mg of (Z)-endoxifen-D-gluconate for 21 days prior to surgery.

The biomarker KI-67 levels of the tumor will be compared from the timeof initial biopsy and the surgical sample will be compared to determineif one of the 3 doses results in a decrease in tumor activity.

Example 18: Therapeutic Treatment of Breast Cancer by OralAdministration of (Z)-Endoxifen D-Gluconate

An objective of the study is to demonstrate that therapeutic(Z)-endoxifen levels in the subject's plasma can be achieved intamoxifen-refractory patients initiating adjuvant breast cancer therapy.Plasma endoxifen levels will be used as a surrogate endpoint to predictclinical benefit as well as the rate of recurrence in this populationand to confirm the safety and tolerability of oral (Z)-endoxifenD-gluconate administration when compared with tamoxifen.

Generally healthy subjects with estrogen receptor positive breast cancerfor whom tamoxifen is indicated as part of their complete treatmentregimen will be enrolled for the study for up to 6 months. At leastseventy five tamoxifen-refractory breast cancer subjects will beenrolled in the study to ensure an evaluable population consists of atleast 25 tamoxifen and 25 (Z)-endoxifen subjects is achieved. Additionalsubjects may be enrolled to achieve the desired population oftamoxifen-refractory subjects. These subjects will have been diagnosedwith breast cancer and undergone mastectomy or lumpectomy.

Immediately after mastectomy or lumpectomy, 25 subjects will be assignedto the tamoxifen group and administered daily with oral 20 mg Tamoxifenfor a period of 3 months. Additional 25 subjects will be assigned to the(Z)-endoxifen group and administered daily with oral 10 mg tablet of(Z)-endoxifen D-gluconate for a period of 3 months. Blood will be drawnfrom each subject at day 0 (baseline), and on days 7, 14, 21, 28, 60 and90. Plasma (Z)-endoxifen levels, hematology, chemistry, and coagulationparameters will be taken for both treatment sets on day 0 (baselinemeasurement), as wells as on days 7, 14, 21, 28, 60 and 90.

The plasma (Z)-endoxifen levels of the two treatment sets will becompared to each other. If the plasma endoxifen levels of the tamoxifentreated subjects are less than 30 nM, then those subjects will betransferred to the (Z)-endoxifen therapy group and the treatment willcontinue for an additional 6 months. If the plasma endoxifen levels ofthe tamoxifen treated subjects are above 30 nM, they will continue to betreated with tamoxifen. Bloods samples will continue to be drawn weeklyuntil the end of the study to monitor the subjects' plasma (Z)-endoxifenlevels, hematology, chemistry, and coagulation parameters. Recurrence ofcancer in the subjects will be monitored and compared between thetreatment groups. Safety and efficacy will be evaluated every 3 monthsuntil the end of the study.

While preferred embodiments of the present invention have been shown anddescribed herein, it will be obvious to those skilled in the art thatsuch embodiments are provided by way of example only. Numerousvariations, changes, and substitutions will now occur to those skilledin the art without departing from the invention. It should be understoodthat various alternatives to the embodiments of the invention describedherein may be employed in practicing the invention. It is intended thatthe following claims define the scope of the invention and that methodsand structures within the scope of these claims and their equivalents becovered thereby.

What is claimed is:
 1. A composition comprising a crystalline form of acompound of Formula (III):

wherein at least 90% by weight of the compound of Formula (III) in thecomposition is the (Z)-isomer, and wherein the crystalline form of the(Z)-isomer is Form I, characterized by an x-ray powder diffractionpattern comprising major peaks at 16.8±0.3°, 17.1±0.3° and 21.8±0.3° twotheta.
 2. The composition of claim 1, wherein the x-ray powderdiffraction pattern further comprises at least one peak selected from16.0±0.3°, 18.8±0.3° and 26.5±0.3 two theta.
 3. The composition of claim1, wherein the x-ray powder diffraction pattern further comprises atleast one peak selected from 12.3±0.3°, 28.0±0.3° and 29.0±0.3° twotheta.
 4. The composition of claim 1, wherein the x-ray powderdiffraction pattern further comprises peaks at 12.3±0.3°, 16.0±0.3°,18.8±0.3°, 26.5±0.3°, 28.0±0.3° and 29.0±0.3° two theta.
 5. Thecomposition of claim 1, wherein the crystalline form of the (Z)-isomeris characterized by an x-ray powder diffraction pattern substantially asset forth in FIG. 9 or FIG.
 10. 6. The composition of claim 1, whereingreater than 90% by weight of the (Z)-isomer in the composition iscrystalline Form I.
 7. The composition of claim 1, further comprising apharmaceutically acceptable carrier or diluent.
 8. The composition ofclaim 7, wherein the composition is formulated for oral, parenteral,topical, or intraductal delivery.
 9. The composition of claim 7, whereinthe composition is formulated for oral delivery as a tablet, a caplet, acapsule, or a pill.
 10. The composition of claim 7, wherein a meanhalf-life of the compound of Formula (III) in a subject treated with thecomposition is between 30 hours to 60 hours.
 11. The composition ofclaim 1, formulated as an oral dosage form comprising 1 mg to 200 mg perunit dose of the composition, and wherein daily administration of theoral dosage form achieves in a subject treated with the composition oneor more of: a steady state plasma level of the compound of Formula (III)within 7 to 21 days; a steady state plasma level of the compound ofFormula (III) ranging from 25 nM to 300 nM; a steady state plasma levelof the compound of Formula (III) greater than 30 nM; or maximal plasmalevels of the compound of Formula (III) within 2 to 10 hours afteradministering.
 12. The composition of claim 11, wherein a mean half-lifeof the compound of Formula (III) in the subject is between 40 hours to55 hours.
 13. The composition of claim 11, wherein the oral dosage formis formulated as an enteric tablet, an enteric caplet, an entericcapsule, a delayed-release tablet, a delayed-release caplet or adelayed-release capsule.
 14. The composition of claim 11, wherein atleast 80% of the compound of Formula (III) in the oral dosage form isreleased in the intestines of the subject.
 15. The composition of claim11, having a mean area under the curve extrapolated to time infinity(AUC_(0-inf)) of the compound of Formula (III) of 200 hr*ng/mL to 10000hr*ng/mL.
 16. A method of treating a hormone-dependent breast disorderor a hormone-dependent reproductive tract disorder in a subject, themethod comprising administering to the subject a composition comprisinga crystalline form of a compound of Formula (III):

wherein at least 90% by weight of the compound of Formula (III) in thecomposition is the (Z)-isomer, and wherein the crystalline form of the(Z)-isomer is Form I, characterized by an x-ray powder diffractionpattern comprising major peaks at 16.8±0.3°, 17.1±0.3° and 21.8±0.3° twotheta, thereby treating the hormone-dependent breast disorder or thehormone-dependent reproductive tract disorder.
 17. The method of claim16, wherein the subject has prostate cancer and wherein the subjectfurther has or is at risk of having gynecomastia.
 18. The method ofclaim 16, wherein the subject has tamoxifen-refractory or tamoxifenresistant hormone-dependent breast disorder or hormone-dependentreproductive tract disorder.
 19. The method of claim 16, wherein thecomposition comprises 0.01 mg to 200 mg of the (Z)-isomer.
 20. Themethod of claim 16, wherein a steady state plasma level of the compoundof Formula (III) is achieved within 7 to 21 days of the firstadministration of the composition to the subject, and wherein the steadystate plasma level of the compound of Formula (III) in the subject isgreater than 30 nM.
 21. The composition of claim 1, wherein thecomposition is stable for at least 9 months at 5° C. and 60% relativehumidity or at 25° C. and 60% relative humidity.